{"paper_id":"2f996d8a-c04e-45c7-8bf3-4394cfb2d482","body_text":"Regeneration of in vitro plants through direct and indirect organogenesis from Dracocephalum rupestre leaf explants | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Regeneration of in vitro plants through direct and indirect organogenesis from Dracocephalum rupestre leaf explants Jia li Wang, Kang jie Yue, Hui xin Liu, Xuping Tian This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4968870/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 25 Mar, 2025 Read the published version in Plant Cell, Tissue and Organ Culture (PCTOC) → Version 1 posted 4 You are reading this latest preprint version Abstract Leaf explants of Dracocephalum rupestre were utilized for regeneration employing direct and indirect differentiation pathways. Results revealed that the direct regeneration medium for leaf explants consisted of MS + 2 mg/l 6-BA + 0.1 mg/l KT + 0.05 mg/l NAA, yielding a differentiation rate of 63.46%. The induction medium for callus was composed of MS + 2 mg/l 6-BA + 0.1 mg/l 2,4-D + 0.5 mg/l IAA, resulting in an induction rate of 86.73%. For the differentiation of adventitious buds, the medium included MS + 2 mg/l 6-BA + 2 mg/l TDZ + 0.05 mg/l IAA, with a differentiation rate of 53.48%. The proliferation medium for adventitious buds generated through both pathways, comprised MS + 2 mg/l 6-BA + 0.05 mg/l NAA, with proliferation rates of 83.57% and 87.41%, respectively. The rooting medium suitable for both methods was 1/2MS + 0.1 mg/l NAA + 0.1 mg/l IBA, resulting in rooting rates of 83.69% and 79.15%, respectively. Comparatively, the direct differentiation pathway proved to be more efficient and time-saving, with leaf explants requiring 30 days less for regeneration compared with the indirect pathway. This study provides theoretical and technical support for subsequent genetic transformation research of Dracocephalum rupestre . Dracocephalum rupestre differentiation pathways callus induction proliferation rooting Figures Figure 1 Figure 2 Introduction Dracocephalum rupestre Hance, also known as rock blue orchid, is a perennial herbaceous plant in Lamiaceae . Its flowers are blue purple in color and have high ornamental values (Yue Zong, 2019). There are various medicinal ingredients such as flavonoids and polysaccharides in D. rupestre , which has been widely used Chinese medicine and have antibacterial, anti-inflammatory, and heat clearing effects; It can also be used to treat symptoms such as cough and inflammation (YuhaoYang, 2023). D. rupestre mainly propagates through seeds, ramets, and bulbils (Tian Xu-ping et al, 2021), these methods of reproduction are inefficient, time-consuming, and easily affected by natural conditions, greatly limiting the development and application of Dracocephalum Rupestre . Therefore, tissue culture has become an important means of its rapid reproduction. Currently, there have been some researches on cultivation of D. rupestre (Liu Jun et al, 2017; Zong Yue, 2019), which mainly focused on different types of explants were used for callus induction, but during the process of callus redifferentiation, only adventitious roots were differentiated, making it difficult to differentiate adventitious buds and a complete regeneration system has not yet been established. This study used the leaves of tissue cultured seedlings of Dracocephalum Rupestre as materials to investigate the effects of different hormones on leaf differentiation pathways, callus induction, bud proliferation, and rooting culture. A complete regeneration system of Dracocephalum Rupestre was established through direct and indirect differentiation pathways, achieving the preservation of its germplasm resources and rapid propagation of seedlings. Materials and Methods Plant materials Leaf blades were harvested from tissue-cultured seedlings of Dracocephalum rupestre after 30 days of cultivation on 1/2 MS medium, with shoots bearing bud segments. The tender leaves at the apex of the plants were selected for the experiment. Direct differentiation pathway The basal culture medium consisted of MS supplemented with 30 g·L − 1 sucrose and 7 g·L − 1 agar. Various concentrations of plant growth regulators were added, including 6-benzylaminopurine(6-BA)at 0.5, 1, and 2 mg/l, kinetin (KT) at 0.1, 0.5, and 1 mg/l, and naphthaleneacetic acid (NAA) at 0.01, 0.05, and 1 mg/l. Each treatment involved inoculating 20 leaf samples with three replicates. The differentiation rate was calculated after 30 days culture. Callus Induction The basal culture medium consisted of MS supplemented with 30 g·L − 1 sucrose and 7 g·L − 1 agar. Different combinations of plant growth regulators were added, including 6-BA at concentrations of 0.5, 1, and 2 mg/l, and a combination of 2,4-dichlorophenoxyacetic acid (2,4-D) and indole-3-acetic acid (IAA) at concentrations of 0.1, 0.5, and 1 mg/l each. Each treatment involved inoculating 20 leaf samples, with three replicates. After 30 days, both the callus induction rate and the growth status of calli were assessed. Adventitious shoot differentiation The basal culture medium comprised MS supplemented with 30 g·L − 1 sucrose and 7 g·L − 1 agar. Various combinations of plant growth regulators were added, including 6-BA at concentrations of 1, 2, and 3 mg/l, thidiazuron (TDZ) at concentrations of 0.5, 1, and 2 mg· −1 , along with combinations of IAA and NAA at concentrations of 0.05, 0.5, and 1 mg/l each. Each treatment involved inoculating 20 pieces of callus tissue, with three replicates. After 30 days, the rates of adventitious shoot and root differentiation, as well as the average number of regenerated shoots, were recorded. Adventitious shoot proliferation The basal culture medium consisted of MS supplemented with 30 g·L − 1 sucrose and 7 g·L − 1 agar. Differentiated adventitious shoots were transferred to media containing combinations of 6-BA at concentrations of 0.5, 1, and 2 mg/l, and NAA at concentrations of 0.01, 0.05, and 0.1 mg/l. Fifteen adventitious shoots were inoculated for each treatment, with three replicates per treatment. After 30 days, the proliferation of adventitious shoots was assessed, and parameters such as proliferation rate, proliferation coefficient, and plant height were measured. Adventitious root formation The basal culture medium comprised 1/2 MS supplemented with 30 g·L − 1 sucrose and 7 g·L − 1 agar. The proliferated adventitious shoots were dissected into single shoots, which were then individually transferred to 16 different media containing various concentrations of NAA (0, 0.1, 0.5, 1 mg/l) or IBA (0, 0.1, 0.5, 1 mg/l). Fifteen shoot explants were inoculated for each treatment, with three replicates per treatment. After 30 days, the rooting of shoots was observed, and rooting percentage as well as plant height were recorded. Hardening and transplantation After removing the tissue-cultured seedlings, approximately 3 cm in height with roots about 2 cm long, they were immersed in distilled water. Following a 2-day placement at room temperature, they were transferred outdoors for a 3-day acclimatization period. Subsequently, they were removed and the culture medium was washed away with distilled water. Finally, they were transplanted into pots containing a 1:1 volume ratio mixture of sterilized vermiculite and nutrient soil. Data collection and analysis Browning rate (%) = Number of browning occurrences / Number of inoculations×100%; Callus induction rate(%) = Number of explants with callus / Number of inoculations×100%; Differentiation rate(%) = Number of calli differentiating into shoots/Number of inoculations× 100%; Shoot proliferation rate (%) = Number of shoots proliferating / Number of inoculations× 100%; Average regenerated shoots (per) = Total number of regenerated shoots / Total number of regenerated explants; Rooting rate (%) = Number of rooted seedlings (or leaves) / Number of surviving seedlings (or leaves) × 100%; Variance analysis and significance analysis were conducted using SPSS 23.0 statistical software. Measurement data were calculated and charts were made using Excel 2016. All data represent the average of three repeated experiments. Results Direct leaf regeneration The leaf began to expand on the 7th day of inoculation (Fig. 1A), with two distinct patterns of adventitious root differentiation observed. One emerged subsequent to leaf browning, characterized by dense and short root systems (Fig. 1B) and the other formed prior to leaf browning, exhibiting longer root systems (Fig. 1C). The leaf browning rate ranged from 26.67–51.34%, while rooting rates from 13.67–72.86%, and budding rates from 0–63.46% (Table 1 ). Significantly higher rooting rates were observed when 6-BA and KT concentrations were set at 0.5 mg/l, and NAA concentration at 1 mg/l, compared to other treatments ( P < 0.05). Notably, leaf browning rates significantly elevated when the KT concentration was higher than the 6-BA concentration (treatment 3) (P < 0.05)No discernible adventitious bud differentiation but higher rooting rates were observed in treatments 1, 2, and 4. Elevated budding rates (Fig. 1D), lower browning rates, and an average regeneration count of 4.39 were revealed in treatment 7. Collectively, The optimal medium for direct leaf regeneration of adventitious buds was determined to be MS + 2 mg/l6-BA + 0.1 mg/lKT + 0.05 mg/lNAA. Table 1 Effects of various hormone combinations on leaf primordia direct differentiation Treatment 6-BA (mg/l) KT (mg/l) NAA (mg/l) Browning rate(%) Rooting rate(%) Germination rate(%) Average regeneration bud count 1 0.5 0.1 0.01 39.34 ± 9.23 b 55.74 ± 10.33 ab - - 2 0.5 0.5 1 27.14 ± 8.52 c 72.86 ± 11.06 a - - 3 0.5 1 0.05 51.34 ± 13.54 a 36.76 ± 13.77 bc 11.76 ± 6.58 c 1.63 ± 0.26 b 4 1 0.1 1 38.81.±8.59 b 61.19 ± 11.76 ab - - 5 1 0.5 0.05 37.08 ± 7.38 b 48.39 ± 10.23 b 16.13 ± 3.58 c 1.76 ± 0.35 b 6 1 1 0.01 40.63 ± 13.22 ab 34.38 ± 7.98 bc 26.56 ± 6.82 bc 1.93 ± 0.21 b 7 2 0.1 0.05 26.67 ± 6.59 c 13.67 ± 9.74 c 63.46 ± 12.38 a 4.39 ± 1.14 a 8 2 0.5 0.01 29.51 ± 9.71 c 49.18 ± 10.56 b 21.31 ± 4.22 bc 3.4 ± 0.74 ab 9 2 1 1 33.87 ± 7.89 bc 37.10 ± 8.69 bc 29.03 ± 9.29 b 2.33 ± 0.42 b Note: Different lowercase letters following data in the same column indicate statistical differences (P < 0.05). “-”: Indicates undifferentiated leaves without adventitious buds Figure 1 The various cultivation stages of direct leaflet differentiation Note: A. 7 days post-inoculation; B. Rooting with browning of leaflets observed 30 days post-inoculation; C. Rooting without browning of leaflets observed 30 days post-inoculation; D. Shoot formation observed 30 days post-inoculation; E. Shoot proliferation; F. Rooting initiation on the 8th day; G. Rooting progression on the 15th day; H. Seedling hardening. Callus Induction The expansion and curling of leaves began on the 7th day after inoculation (Fig. 2A), followed by the induction of callus 15 days later (Fig. 2B). Table 2 reveals that the induction rate of callus from leaves ranged between 19.03% and 86.73% across different hormone combinations in the culture medium. With consistent 6-BA concentrations, the callus induction rate gradually decreased with increasing concentrations of 2,4-D and IAA. At 0.5 and 2 mg/l of 6-BA, the induction rate was higher with the sole addition of 2,4-D compared to IAA, while at 1 mg/l IAA, the induction rate was higher than that with 2,4-D. Among the media containing 6-BA, 2,4-D, and IAA, Treatment 25 exhibited the highest induction rate at 86.73%. The resulting callus appeared yellowish-green and loose. In summary, Treatment 25, with MS + 2 mg/l 6-BA + 0.1 mg/l 2,4-D + 0.5 mg/l IAA, was determined to be the optimal culture medium for inducing callus from leaves. Table 2 Effects of different hormone ratios on leaf callus induction Treatment 6-BA (mg/l) 2,4-D (mg/l) IAA (mg/l) Induction rate (%) Callus growth state 1 0.5 0.1 - 44.17 ± 9.31 c Pale yellow, loose, soft texture 2 0.5 0.5 - 39.98 ± 8.74 cd Pale yellow, loose, hard texture 3 0.5 1 - 19.03 ± 6.33 d Yellow, loose, soft texture 4 1 0.1 - 49.71 ± 10.63 bc Green loose, softer texture 5 1 0.5 - 44.54 ± 9.63 c Yellow, compact, hard texture 6 1 1 - 38.05 ± 8.94 cd Pale yellow, loose, softer texture 7 2 0.1 - 66.11 ± 11.82 b Green loose, softer texture 8 2 0.5 - 53.42 ± 11.44 bc Pale yellow, loose, softer texture 9 2 1 - 50.76 ± 14.13 bc Pale yellow, loose, softer texture 10 0.5 - 0.1 34.52 ± 6.34 cd Yellow-green, loose,soft texture 11 0.5 - 0.5 31.96 ± 9.75 cd Yellow-green, compact,softer texture 12 0.5 - 1 24.07 ± 7.22 cd Yellow-green, loose,soft texture 13 1 - 0.1 51.34 ± 11.99 bc Yellow-green, loose, soft texture 14 1 - 0.5 46.21 ± 11.23 c Yellow-green loose softer texture 15 1 - 1 39.14 ± 9.41 cd Yellow-green loose, soft texture 16 2 - 0.1 55.24 ± 6.91 bc Yellow-green loose, soft texture 17 2 - 0.5 51.33 ± 13.54 bc Green loose, softer texture 18 2 - 1 46.61 ± 14.26 c Yellow-green loose, softer texture 19 0.5 0.1 0.1 47.62 ± 9.43 c Green, compact, hard texture 20 0.5 0.5 1 53.41 ± 13.22 bc Green, compact, hard texture 21 0.5 1 0.5 55.76 ± 10.16 bc Green, compact, somewhat hard texture 22 1 0.1 1 42.57 ± 9.25 c Yellow-green loose, softer texture 23 1 0.5 0.5 50.91 ± 13.22 bc Pale yellow, loose, softer texture 24 1 1 0.1 44.59 ± 11.17 c Yellow-green,loose, softer texture 25 2 0.1 0.5 86.73 ± 11.29 a Yellow-green,loose, softer texture 26 2 0.5 0.1 70.23 ± 17.05 ab Yellow loose, soft texture 27 2 1 1 67.15 ± 13.21 b Green compact,hard texture Callus Differentiation of leaf After 30 days post-inoculation, callus differentiated into adventitious roots or shoots (Fig. 2C, D). As shown in Table 3 , callus failed to differentiate into shoots when solely supplemented with either 6-BA or TDZ. The rooting rate decreased with increasing concentrations of 6-BA, while it increased with TDZ concentrations, although the differences were not significant. Simultaneous addition of both 6-BA and TDZ yielded shoot differentiation rates ranging from 0–7.66%, and rooting rates ranging from 0–9.17%. Treatments involving the concurrent addition of 6-BA, TDZ, and NAA resulted in only treatments 16, 17, and 19 differentiating into adventitious roots, while the remaining six treatments exhibited diverse outcomes, with some producing adventitious roots and others generating shoots. Treatment 21 exhibited the highest shoot differentiation rate at 53.48%, while treatment 17 had the highest rooting rate at 23.64%. On average, treatment 21 yielded the highest number of adventitious shoots at 4.23 per explant. Overall, the optimal callus differentiation medium for shoot induction was treatment 21, consisting of MS supplemented with 2 mg/l 6-BA, 2 mg/l TDZ, and 0.5 mg/l IAA. Table 3 Effects of different hormone ratios on leaf callus differentiation Treatment 6-BA (mg/l) TDZ (mg/l) IAA (mg/l) Germination rate(%) Rooting rate (%) Average number of adventitious buds 1 1 0 - - 4.58 ± 1.51 c - 2 2 0 - - 3.43 ± 1.02 cd - 3 3 0 - - 2.33 ± 0.93 cd 1.26 ± 0.29c 4 0 0.5 - - - - 5 0 1 - - 4.52 ± 0.95 c - 6 0 2 - - 4.74 ± 1.39 c - 7 1 0.5 - - 1.92 ± 0.82 d 1.22 ± 0.20 c 8 1 1 - - 9.17 ± 2.42 bc - 9 1 2 - - 6.11 ± 2.21 c - 10 2 0.5 - 3.44 ± 1.32 c 8.32 ± 2.31 bc 1.56 ± 0.33 c 11 2 1 - - 6.41 ± 2.13 c 2.03 ± 0.43 bc 12 2 2 - 2.91 ± 0.71 c 3.59 ± 1.33 c - 13 3 0.5 - 2.37 ± 0.69 c 7.15 ± 1.12 bc 3.08 ± 0.93 b 14 3 1 - 3.17 ± 1.21 c - - 15 3 2 - 7.66 ± 2.12 c - 1.41 ± 0.22 c 16 1 0.5 0.05 - 14.32 ± 4.23 b - 17 1 1 1 - 23.64 ± 9.34 a - 18 1 2 0.5 22.35 ± 9.33 b 4.52 ± 1.96 c 3.27 ± 0.53 b 19 2 0.5 1 - 17.61 ± 3.22 ab - 20 2 1 0.5 1.32 ± 0.42 c 9.02 ± 2.83 bc 1.93 ± 0.33 bc 21 2 2 0.05 53.48 ± 9.89 a 6.49 ± 2.02 c 4.23 ± 1.06 a 22 3 0.5 0.5 9.46 ± 2.31 c 19.03 ± 7.93 ab 4.06 ± 0.95 a 23 3 1 0.05 19.83 ± 6.99 b 12.87 ± 4.85 b 3.61 ± 0.84 b 24 3 2 1 9.10 ± 2.41 c 8.99 ± 3.31 bc 2.77 ± 0.32 bc Adventitious Shoot Proliferation Following grafting of adventitious buds onto nine types of bud proliferation culture media, the bud volume increased, and growth rate accelerated. After 20 days, adventitious buds proliferated around the bud blocks. The proliferation rates of adventitious buds via direct differentiation ranged from 7.53–87.41%, while those via indirect differentiation ranged from 17.21–83.57%(Table 4 ). At a concentration of 0.5 mg/l 6-BA, both types of buds exhibited decreased proliferation rates with increasing NAA concentration. Notably, adventitious buds regenerated through the indirect pathway displayed higher proliferation rates. At 1 mg/l, the proliferation rate of adventitious buds via direct differentiation initially increased and then decreased with rising NAA concentration, while the rate via indirect differentiation gradually increased. At 2 mg/l, the proliferation rate of adventitious buds via direct regeneration initially increased and then decreased, whereas the rate via indirect differentiation gradually decreased. The highest proliferation rates for adventitious buds were 87.41% (Fig. 1E) and 82.96% (Fig. 2E), with proliferation coefficients exceeding 4-fold and 3-fold, respectively. Overall, the proliferation rate of adventitious buds via direct regeneration surpassed that via indirect regeneration. Treatment 8, comprising MS + 2 mg/l 6-BA + 0.05 mg/l NAA, proved suitable for direct regeneration of adventitious buds, while treatment 7, consisting of MS + 2 mg/l 6-BA and 0.01 mg/l NAA, was deemed suitable for indirect regeneration of adventitious buds. Table 4 Effects of different hormone combinations on adventitious bud proliferation Direct regeneration Indirect regeneration Treatment 6-BA (mg/l) NAA (mg/l) Bud growth rate (%) Coefficient of increment Bud growth rate (%) Coefficient of increment 1 0.5 0.01 22.53 ± 7.95 c + 56.85 ± 6.94 bc + 2 0.5 0.05 16.73 ± 3.94 c + 37.16 ± 6.77 c + 3 0.5 0.1 7.53 ± 1.21 c + 19.81 ± 4.31 c + 4 1 0.01 69.31 ± 10.97 b +++ 65.36 ± 12.74 b +++ 5 1 0.05 73.86 ± 10.47 ab +++ 70.39 ± 11.91 ab ++ 6 1 0.1 61.91 ± 9.85 b ++ 76.59 ± 14.61 ab ++ 7 2 0.01 81.69 ± 13.61 a ++++ 82.96 ± 11.85 a +++ 8 2 0.05 87.41 ± 12.56 a ++++ 74.27 ± 9.81 ab +++ 9 2 0.1 82.39 ± 15.31 a ++ 71.51 ± 14.33 ab +++ ++++ means the multiplication factor is more than 4 time +++ means the multiplication factor is more than 3 times; ++ means the multiplication factor is about 2 times; + means the multiplication factor is about 1 times; Adventitious Shoot Rooting After transferring well-developed seedlings to the rooting medium, root systems formed on the 8th day (Fig. 1F, Fig. 2F), and the roots gradually elongated after 15 days (Fig. 1G, Fig. 2G). As shown in Table 5 , among the 10 rooting media, the rooting rate of direct regeneration adventitious buds ranged from 46.17–83.69%, while that of indirect regeneration adventitious buds ranged from 43.67–79.63%. The rooting rates of adventitious buds for both differentiation pathways increased with higher NAA concentrations and initially increased then decreased with higher IBA concentrations. At the same concentration, the rooting rate with NAA was better than with IBA. When NAA and IBA were combined at both concentrations of 0.1 mg·L − 1 , the highest rooting rates were achieved, 83.69% and 79.63%, respectively, with a decrease in rooting rate as the concentration increased.For the direct regeneration pathway, the plant height of adventitious buds increased with higher NAA and IBA concentrations, while for the indirect regeneration pathway, the plant height first increased then decreased with higher concentrations of NAA and IBA. When NAA and IBA concentrations were both at 0.1 mg·L − 1 , the plant heights of adventitious buds for the two differentiation pathways were 4.52 cm and 3.79 cm, respectively, significantly higher than other treatments ( P < 0.05).In summary, NAA has a better effect on adventitious bud rooting of Mimosa pudica than IBA, and the direct regeneration pathway is more effective than the indirect regeneration pathway. The optimal rooting medium for adventitious buds is treatment 8, 1/2 MS + 0.1 mg·L − 1 NAA + 0.1 mg·L − 1 IBA. After acclimatization, the plants were transplanted into a substrate mixed with perlite and potting soil in a 1:1 ratio, and the plants grew robustly (Fig. 1H, Fig. 2H). Table 5 Effects of different concentrations of auxins on adventitious bud rooting Direct regeneration Indirect regeneration Treatment NAA (mg/l) IBA (mg/l) Rooting rate(%) Plant height(cm) Rooting rate(%) Plant height(cm) 1 0 - 56.93 ± 10.51 bc 2.63 ± 0.32 bc 51.21 ± 9.52 bc 2.63 ± 0.32 b 2 0.1 - 67.39 ± 12.79 b 2.86 ± 0.71 bc 57.83 ± 11.54 b 1.62 ± 0.29 c 3 0.5 - 83.69 ± 13.49 a 3.97 ± 0.85 b 60.36 ± 9.81 b 2..21 ± 0.70 bc 4 1 - 73.55.±8.56 a 1.67 ± 0.41 c 47.69.±11.32 c 2.42 ± 0.59 bc 5 - 0.1 46.73 ± 10.54 c 2.32 ± 0.40 bc 73.27 ± 14.13 ab 1.44 ± 0.13 c 6 - 0.5 59.71 ± 12.17 bc 2.94 ± 0.77 bc 70.26 ± 8.17 b 1.87 ± 0.55 c 7 - 1 63.19 ± 9.85 b 3.09 ± 0.49 b 51.82 ± 13.62 c 2.53 ± 0.37 b 8 0.1 0.1 74.28 ± 11.19 a 4.03 ± 0.93 a 80.15 ± 12.38 a 3.89 ± 0.66 a 9 0.5 0.5 51.66 ± 8.41 bc 3.01 ± 0.52 b 71.41 ± 11.46 ab 3.55 ± 0.46 ab 10 1 1 46.17 ± 13.22 c 1.36 ± 0.34 c 51.11 ± 8.71 c 2.34 ± 0.27 bc Figure 2 The various cultivation stages of the indirect leaflet differentiation pathway Note: A. Induction of callus tissue occurs 7 days after inoculation; B. Callus tissue induction is observed 15 days post-inoculation; C. Adventitious roots emerge from the callus tissue 30 days post-inoculation; D. Adventitious shoots develop from the callus tissue 30 days post-inoculation. E. Proliferation of adventitious shoots; F. Rooting begins on the 8th day; G. Rooting progresses by the 15th day; H. Seedling hardening. Discussion Plant regeneration includes two differentiation methods: direct regeneration and indirect regeneration (Long Y et al, 2022). The direct differentiation pathway is the process of selecting explants that do not undergo callus induction and directly differentiate into adventitious buds to obtain regenerated plants; The indirect differentiation pathway involves the induction of callus tissue, adventitious bud differentiation, and rooting of explants to obtain regenerated plants (Jiang W et al, 2012). Direct regeneration path the diameter can obtain regenerated plants in a short period of time, which has the advantages of high efficiency and time saving; The indirect differentiation pathway iswidely used, but the cultivation cycle is long and the workload is large(Youssef N M et al 2019). In Lamiaceae , indirect regeneration pathways are commonly used, with a cultivation period of over 120 days (Liu J H et al 2022), however, there is relatively little research on direct regeneration. In this study, the regeneration of the indirect differentiation pathway using the leaves of Dracocephalum rupestre pubescens as explants took 120 days, which is similar to the previous research findings. The direct differentiation pathway took only 90 days to obtain regenerated plants, At NAA and IBA concentrations of 0.1 mg/l, the rooting rate and plant height of shoots obtained through direct differentiation pathways are superior to those through indirect pathways. Moreover, compared to the indirect pathway, the direct pathway exhibited stronger genetic stability with no variation in genotype and phenotype traits, making it suitable for regenerating transgenic plants (Hafez et al, 2013 ). Lamiaceae tissue culture commonly uses certain concentrations of cytokinins (6-BA, TDZ, KT)and auxin (NAA, IAA, etc.)2.4-D induced callus and differentiation (Kosar et al 2012, Gharari Z, 2021). The color and quality ofcallus tissue are influenced by both. The impact of concentration, 6-BA and 2, 4-D) combination suitable for Lavandula angustifolia (Chan Wang et al, 2012). Leaf callus induction, when the ratio of the two is 1:l, results in better callusquality and stronger differentiation ability in the later stage. The concentration of 6-BA and NAA at 10:1 is suitable for inducing callus tissue from Dracocephalum tanguticum leaves. In previous studies, a ratio of 6-BA to NAA at 1:1 can induce callus formation in the leaves of Phyllostachys sinensis, but its color is yellow and itstexture is loose. In the later stage, only adventitious roots are differentiated, and its ability to further produce adventitious buds is poor (Yue Zong, 2019). In this study, the induction rate and quality of leaf callus tissue showed different states in different hormone combinations. In the combination of 6-BA and asingle combination of 2,4-D or IAA, the induction rate of leaf callus tissue is low, and the callus tissue is in poor condition, which is not conducive to the differentiation of adventitious buds in the later stage. However, in the medium with the addition of 6-BA, 2,4-D and IAA, the induction rate of callus tissue is the highest at 86. 73%. At this time, the callus tissue is loose and the color is yellow green. This is different from previous research results, possibly because the synergistic effect of multiplehormones is much greater than that of a single hormone used alone, and its effect is closely related to the type and concentration of endogenous hormones in the explant andcallus tissue itself (Darvishi et al, 2014; Gharari et al, 2021 ). During the differentiation stage, the mesophyll cells of explants possess dual characteristics of high auxin accumulation and hypersensitivity to cytokinins, enabling the explants to regenerate both roots and shoots. When the medium contains high concentrations of auxin and low levels of cytokinins, the explants differentiate into roots due to the accumulated auxin. Conversely, high concentrations of cytokinins in the medium exploit the hypersensitivity of callus tissues, rapidly and effectively activating shoot gene expression, thereby promoting shoot development (Bidabadi et al 2020). In the direct regeneration of leaves in this study, when NAA concentrations were at 1 mg/l and 6-BA or KT concentrations were at 0.5 mg/l or 0.1 mg/l, a higher rooting rate was observed with no adventitious bud differentiation. Conversely, higher concentrations of 6-BA at 2 mg/l and lower concentrations of NAA at 0.01–0.05 mg/l resulted in higher adventitious bud differentiation rates but lower rooting rates. In the indirect regeneration of leaves, higher IAA concentrations at 1 mg/l with 6-BA or TDZ at 1 mg/l led to higher rooting rates but lower adventitious differentiation rates, consistent with the aforementioned research findings. For Dracocephalum rupestre leaf explants, higher rates of adventitious root differentiation were observed in media containing combinations of 6-BA and NAA, with no adventitious bud formation (Zong Y, 2019), while IAA facilitated bud induction in Dracocephalum rupestre (Liu J et al, 2017). In this study, the induction rates of adventitious buds in callus tissues were significantly lower with the addition of single or combined 6-BA and TDZ, whereas the addition of IAA markedly enhanced the differentiation rate of adventitious buds, aligning with previous research conclusions (Liu J et al, 2017). This suggests a close relationship between the regeneration of adventitious roots or buds in explants and the types of exogenous hormones used. The high rate of adventitious root differentiation and low rate of adventitious bud differentiation in direct or indirect regeneration processes of Dracocephalum rupestre leaf explants may also be related to the content of endogenous hormones within the plant (Gharari et al, 2021 ). However, further investigation is required to elucidate the specific changes in endogenous hormone types and levels. The combination of 6-BA and NAA is commonly used in the propagation of Labiatae species through adventitious bud culture. Optimal bud proliferation occurs when the concentration of 6-BA ranges from 1 to 2 mg/l and NAA ranges from 0.01 to 0.5 mg/l (Xi Y K et al, 2020 ). At 1 mg/l of 6-BA, the proliferation efficiency of direct mint buds decreases with increasing NAA concentration, whereas at 0.5 mg/l, the efficiency of indirect mint bud proliferation increases with higher NAA concentrations (Islam A et al, 2018). 6-BA plays a crucial role in the proliferation of indirect buds in Salvia splendens , but excessive concentrations inhibit bud cluster formation. In media supplemented solely with NAA or IBA, splendens exhibits low rates of adventitious bud proliferation and weaker bud growth (Liu Hui et al, 2011 ). In this study, at a concentration of 0.5 mg/l of 6-BA, the proliferation rates of both differentiation pathways of adventitious buds increase with higher NAA concentrations. At 1 mg/l, the proliferation of direct pathway buds decreases with increasing NAA, while the indirect pathway shows an opposite trend, consistent with previous findings (Islam A et al, 2018). This indicates that different explant sources treated with the same hormone combination exhibit varying proliferation rates. Selecting appropriate combinations of 6-BA and NAA can yield higher proliferation rates and coefficients (Ghasemiomran et al, 2018). Auxin NAA and lBA are commnly used in root culture of plants in the family Lamiaceae , but the concentration of auxin suitable for plant rooting varies(Aghazadeh et al, 2015; Bassolino L et al, 2015). The rooting effect of IBA at the same concentration on Elsholtzia bodinieri is better than that of NAA. As the concentration of IBA increases, the rooting rate of Elsholtzia bodinieri (Zhao Jie et al, 2023 ) increases with the increase of IBA concentration, The appropriate concentration for its rooting is l mg/l. The rooting effect of NAA at the same concentration on Salvia hispanica is better than that of IBA(Li Wei-li et al, 2016). The single addition of NAA and IBA is not conducive to the induction of the root system of Clerodendranthus spicatus (Li Jia-hui et al, 2022). At the same time, adding l mg/l of NAA and IBA each resulted in a higher rooting rate and robust plant growth, In this study, it was found that the hormone combination added with NAA had a better overall rooting effect than the combination added with IBA. When NAA and IBA were added simultaneously, the rooting rate was higher and the rooting rate of adventitious buds increased with the concentration of NAA and IBA. Conclusion In summary, this study established a regeneration system through direct and indirect differentiation pathways using the leaves of tissue cultured seedlings of Dracocephalum rupestre as explants. Research has found that the culture medium for direct regeneration of leaves is MS + 2 mg/l 6-BA + 0.1 mg/l KT + 0.05 mg/l NAA. The differentiation rate of adventitious buds is 63.46%, The callus induction medium is MS + 2 mg/l6-BA + 0.1 mg/l 2,4-D + 0.5 mg/l IAA, with an induction rate of 86.73%. The differentiation medium is MS + 2 mg/l 6-BA + 2 mg/l TDZ + 0.05 mg/l IAA, with an adventitious bud differentiation rate of 66.37%.The adventitious bud proliferation medium for the two differentiation pathways is the same. MS + 2 mg/l 6-BA + 0.05 mg/l NAA, The proliferation rates for direct differentiation and indirect differentiation pathways were 83.57% and 87.41%, respectively. The optimal rooting medium consisted of 1/2 MS supplemented with 0.1 mg/l NAA and 0.1 mg/l IBA, resulting in rooting rates of 83.69% for direct differentiation and 79.15% for indirect differentiation pathways. The direct differentiation pathway takes 90 days from leaf isolation to plant regeneration, which is 30 days less than the indirect differentiation pathway. The process is simple and the differentiation rate, proliferation rate, and rooting rate are higher than the indirect differentiation pathway. This provides the oretical and technical support for the subsequent research on the genetic transformation system of Dracocephalum rupestre . References Aghazadeh P, Behboodi B S(2015)Seed germination and in vitro organogenesis optimization of Zhumeria majdae (Lamiaceae) from Iran. American Journal of Plant Sciences, 6(11): 1850-1856.https//:doi: 10.4236/ajps.2015.611185 Bassolino L, Giacomelli E, Giovanelli S(2015)Tissue culture and aromatic profile in Salvia dolomitica Codd. Plant Cell, Tissue and Organ Culture (PCTOC), 121: 83-95. https//:doi:10.1007/s11240-014-0681-3 Bidabadi S S, Jain S M(2020)Cellular, molecular, and physiological aspects of in vitro plant regeneration. Plants, 9(6): 702. https://doi.org/10.3390/plants9060702 Deepa A V, Thomas D T(2022). Tissue Culture Studies in Lamiaceae: A Review. Biotechnology and Crop Improvement, 181-211. https://doi:10.1201/9781003239932-11 Darvishi E, Kazemi E, Kahrizi D(2014)Optimization of callus induction in Pennyroyal (Mentha pulegium). Journal of Applied Biotechnology Reports, 1(3): 97-100. https://doi:10.1023/A:1000304922507. Gharari Z, Bagheri K, Karimkhanlooei G(2021)Study of tissue culture and in vitro organogenesis of Scutellaria bornmuelleri using benzylaminopurine, lsopentenyl adenine and thidiazuron. South African Journal of Botany, 139(13): 458-469. https://doi.org/10.1016/j.sajb.2021.03.030 Ghasemiomran V, Movahedi M, Alizadeh F(2018)Effects of explant type and plant growth regulator combinations on in vitro direct and indirect regeneration of Stevia rebaudiana.19(23):137-147. https://doi:10.1007/s11627-014-9640-2 Hafez R F, Shahabzadeh Z, Heidari B(2013)Investigation of the efficiency of direct and indirect regeneration in evening primrose (Oenothera biennis). Journal of crop science and biotechnology, 16(09): 291-296. htttps://:doi:10.1007/s12892-013-0115-5 Islam A, Alam M F(2018)In vitro callus induction and indirect organogenesis of Mentha piperita (L.)-an aromatic medicinal plant. GSC Biological and Pharmaceutical Sciences, 4(3): 049-060. https://doi.org/10.30574/gscbps.2018.4.3.0078 Jiang W, Chen L, Pan Q(2012)An efficient regeneration system via direct and indirect organogenesis for the medicinal plant Dysosma versipellis (Hance) M. Cheng and its potential as a podophyllotoxin source. Acta Physiologiae Plantarum,34(23): 631-639. https://doi:10.1007/s11738-011-0864-z Kosar M, Mahmoud O(2012)Trichomes and regeneration by direct organogenesis of medicinal plant Dracocephalum kotschyi L. using shoot tips (Lamiaceae). Journal of Crop Science and Biotechnology, 15(3): 251-257. https://doi:10.1007/s12892-012-0019-9 Long Y, Yang Y, Pan G(2022)New insights into tissue culture plant-regeneration mechanisms. Frontiers in plant science,13(08):752-926. https://doi:10.3389/fpls.2022.926752 Liu J H, Dong W C, Fei F F( 2022)Regulation of WOX11 expression represents the difference between direct and indirect shoot regeneration. Frontiers in Plant Science, 13: 850726. https://doi.org/10.3389/fpls.2022.850726 Liu Hui, Chen Yi, Zhao Hang-ping(2011). Elementary study on the high effective regeneration system of Salvia Splendens. Acta Agriculturae Zhejiangensis. 23(02):318-323. https://doi:10.3969/j.issn.1004-1524.2011.02.024. Liu Jun, Sun Rui-fen, Geng Mu-dan. et al(2017). Establishment of rapid propagation system of Dracocephalum rupestre Hance in vitro culture. Journal of Northern Agriculture, 45(06): 102-106. https://doi:10.3969/j.issn.2096-1197.2017.06.19. Li Wei-li, Lifang Huang, Xinjie Xia(2016)Tissue culture and rapid propagation of Salvia hispanica. Pratacural Science, 33(03):393-399. https:// doi:10.11829/j.issn.1001-0629.2015-0424 Li Jia-hui, Ye Wei-yan, Zhu Peng-jin, et al(2022)Establishment of a sterile short shoot tissue culture and rapid propagation system for Clerodendranthus spicatus. Chinese Journal of Tropical Crops, 43(10): 2063-2070. http://doi: 10.3969/j.issn.1000-2561.2022.10.012 Tian Xu-ping, Guo Qing-hao, Sun Ming-yu, et al(2021). Comparison on bulbil formation of Dracocephalum rupestre Hance originated from different provenances. Acta Agrestia Sinica, 29(06):1357 - 1362. https://doi:10.11733/j.issn.1007-0435.2021.06.028. Wang Chan, Chen Li-juan, Cheng Ming-hua, et al(2012). Study on in vitro cultivation of lavandula angustifolia , Journal of Hainan Normal University（Natural Science）, 25(04): 435–437. https://doi:10.3969/j.issn.1674-4942.2012.04.022. Xi Y K, Wang Y, Zeng B(2020). Callus induction and adventitious bud differentiation of Cyclocodon lancifolius (Roxb.) Kurz. Botanical Sciences, 98(4): 534-544. https://doi:10.17129/botsci.2609 Youssef N M, Shaaban S A, Ghareeb Z F(2019). In vitro bulb formation of direct and indirect regeneration of Lilium orientalis cv.“Starfighter” plants. Bulletin of the National Research Centre, 43: 1-9.https://doi.org/10.1186/s42269-019-0246-z Zong Yue, Guo Qing-hao, Tian Xu-ping(2019). Callus Induction and Subculture Test of Stem Segment of Dracocephalum rupestre. Journal of Shanxi Agricultural Sciences. 47(04):507-509. https:// doi:10.3969/j.issn.1002-2481.2019.04.04. Zhang Jing, Niu Zhe, Fan Wei-fang (2020). Establishing tissue culture and regeneration system of Scutellaria regeliana. Bulletin of Botanical Research, 40(1): 50-59. https://doi:10.7525/j.issn.1673-5102.2020.01.008 Zhao Jie, Hou Yu-qi, Zhen Wei, Xu Xiao-dan(2023). Study on invitro rapid propagation of Elsholtzia bodinieri. Northern Horticulture, 45(23):60-67. https://doi:10.11937/bfyy.20231147 Cite Share Download PDF Status: Published Journal Publication published 25 Mar, 2025 Read the published version in Plant Cell, Tissue and Organ Culture (PCTOC) → Version 1 posted Reviewers agreed at journal 15 Nov, 2024 Reviewers invited by journal 14 Nov, 2024 Editor assigned by journal 21 Oct, 2024 First submitted to journal 21 Oct, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {\"props\":{\"pageProps\":{\"initialData\":{\"identity\":\"rs-4968870\",\"acceptedTermsAndConditions\":true,\"allowDirectSubmit\":false,\"archivedVersions\":[],\"articleType\":\"Research Article\",\"associatedPublications\":[],\"authors\":[{\"id\":378125439,\"identity\":\"1363566a-0ab8-4b89-911e-3cfe8c683d1e\",\"order_by\":0,\"name\":\"Jia li Wang\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Shanxi Agricultural University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Jia\",\"middleName\":\"li\",\"lastName\":\"Wang\",\"suffix\":\"\"},{\"id\":378125440,\"identity\":\"afc4a07a-f609-40c0-8c54-a951b2b091de\",\"order_by\":1,\"name\":\"Kang jie Yue\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Shanxi Agricultural University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Kang\",\"middleName\":\"jie\",\"lastName\":\"Yue\",\"suffix\":\"\"},{\"id\":378125441,\"identity\":\"86c8eaf9-69a7-4e35-8613-6da5ed428b34\",\"order_by\":2,\"name\":\"Hui xin Liu\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Shanxi Agricultural University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Hui\",\"middleName\":\"xin\",\"lastName\":\"Liu\",\"suffix\":\"\"},{\"id\":378125442,\"identity\":\"84f55eea-01dd-4caf-8070-5679cac0c7e0\",\"order_by\":3,\"name\":\"Xuping Tian\",\"email\":\"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAyklEQVRIiWNgGAWjYBACxvbmg4///mFjZmNvIFILc8+xZAPeBj52Pp4DRGphn5GjJsHbIMcvJ5FApBbeGTkMEpI7zKTZJB9vvMFQYxNNUItkz9sDBoZn0ozZpNOKLRiOpeU2ENJi2J6XkJDAdiyZTTrHTIKx4TBhLfYHcgwOHGD7X98meYZILYwdOYaNjW3AQJbgIVYLMJCZGc4AtfAA/ZJAjF+AUXn8N0MFG7N8++GNNz7U2BDWggwMiI4aJC2k6hgFo2AUjIKRAQBz4zzhM286qAAAAABJRU5ErkJggg==\",\"orcid\":\"https://orcid.org/0009-0001-4904-4763\",\"institution\":\"Shanxi Agricultural University\",\"correspondingAuthor\":true,\"prefix\":\"\",\"firstName\":\"Xuping\",\"middleName\":\"\",\"lastName\":\"Tian\",\"suffix\":\"\"}],\"badges\":[],\"createdAt\":\"2024-08-24 11:16:04\",\"currentVersionCode\":1,\"declarations\":\"\",\"doi\":\"10.21203/rs.3.rs-4968870/v1\",\"doiUrl\":\"https://doi.org/10.21203/rs.3.rs-4968870/v1\",\"draftVersion\":[],\"editorialEvents\":[{\"content\":\"https://doi.org/10.1007/s11240-025-03026-1\",\"type\":\"published\",\"date\":\"2025-03-25T15:57:10+00:00\"}],\"editorialNote\":\"\",\"failedWorkflow\":false,\"files\":[{\"id\":70357538,\"identity\":\"1e3df46a-00d0-4924-94ca-abeaaaaafe69\",\"added_by\":\"auto\",\"created_at\":\"2024-12-02 12:54:52\",\"extension\":\"png\",\"order_by\":1,\"title\":\"Figure 1\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":1567560,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eThe various cultivation stages of direct leaflet differentiation\\u003c/p\\u003e\\n\\u003cp\\u003eNote: A. 7 days post-inoculation; B. Rooting with browning of leaflets observed 30 days post-inoculation; C. Rooting without browning of leaflets observed 30 days post-inoculation; D. Shoot formation observed 30 days post-inoculation; E. Shoot proliferation; F. Rooting initiation on the 8th day; G. Rooting progression on the 15th day; H. Seedling hardening.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage1.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-4968870/v1/3e139f3a59585f9d007d7df9.png\"},{\"id\":70357536,\"identity\":\"ddf1831f-b3d0-4e33-942b-dfa1b6be522d\",\"added_by\":\"auto\",\"created_at\":\"2024-12-02 12:54:52\",\"extension\":\"png\",\"order_by\":2,\"title\":\"Figure 2\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":274098,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eThe various cultivation stages of the indirect leaflet differentiation pathway\\u003c/p\\u003e\\n\\u003cp\\u003eNote: A. Induction of callus tissue occurs 7 days after inoculation; B. Callus tissue induction is observed 15 days post-inoculation; C. Adventitious roots emerge from the callus tissue 30 days post-inoculation; D. Adventitious shoots develop from the callus tissue 30 days post-inoculation. E. Proliferation of adventitious shoots; F. Rooting begins on the 8th day; G. Rooting progresses by the 15th day; H. Seedling hardening.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage2.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-4968870/v1/fddd9e6f7111ed9d973c2248.png\"},{\"id\":79605081,\"identity\":\"af05b828-6153-463e-8185-5d46f344cfb4\",\"added_by\":\"auto\",\"created_at\":\"2025-03-31 16:10:32\",\"extension\":\"pdf\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"manuscript-pdf\",\"size\":2874853,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"manuscript.pdf\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-4968870/v1/66cf3b80-0d22-4a9b-bc3f-bc3d484696a2.pdf\"}],\"financialInterests\":\"\",\"formattedTitle\":\"Regeneration of in vitro plants through direct and indirect organogenesis from Dracocephalum rupestre leaf explants\",\"fulltext\":[{\"header\":\"Introduction\",\"content\":\"\\u003cp\\u003e \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e Hance, also known as rock blue orchid, is a perennial herbaceous plant in \\u003cem\\u003eLamiaceae\\u003c/em\\u003e. Its flowers are blue purple in color and have high ornamental values (Yue Zong, 2019). There are various medicinal ingredients such as flavonoids and polysaccharides in \\u003cem\\u003eD. rupestre\\u003c/em\\u003e, which has been widely used Chinese medicine and have antibacterial, anti-inflammatory, and heat clearing effects; It can also be used to treat symptoms such as cough and inflammation (YuhaoYang, 2023). \\u003cem\\u003eD. rupestre\\u003c/em\\u003e mainly propagates through seeds, ramets, and bulbils (Tian Xu-ping et al, 2021), these methods of reproduction are inefficient, time-consuming, and easily affected by natural conditions, greatly limiting the development and application of \\u003cem\\u003eDracocephalum Rupestre\\u003c/em\\u003e. Therefore, tissue culture has become an important means of its rapid reproduction. Currently, there have been some researches on cultivation of D. rupestre (Liu Jun et al, 2017; Zong Yue, 2019), which mainly focused on different types of explants were used for callus induction, but during the process of callus redifferentiation, only adventitious roots were differentiated, making it difficult to differentiate adventitious buds and a complete regeneration system has not yet been established.\\u003c/p\\u003e \\u003cp\\u003eThis study used the leaves of tissue cultured seedlings of \\u003cem\\u003eDracocephalum Rupestre\\u003c/em\\u003e as materials to investigate the effects of different hormones on leaf differentiation pathways, callus induction, bud proliferation, and rooting culture. A complete regeneration system of \\u003cem\\u003eDracocephalum Rupestre\\u003c/em\\u003e was established through direct and indirect differentiation pathways, achieving the preservation of its germplasm resources and rapid propagation of seedlings.\\u003c/p\\u003e\"},{\"header\":\"Materials and Methods\",\"content\":\"\\u003cdiv id=\\\"Sec3\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003ePlant materials\\u003c/h2\\u003e \\u003cp\\u003eLeaf blades were harvested from tissue-cultured seedlings of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e after 30 days of cultivation on 1/2 MS medium, with shoots bearing bud segments. The tender leaves at the apex of the plants were selected for the experiment.\\u003c/p\\u003e \\u003c/div\\u003e\\n\\u003ch3\\u003eDirect differentiation pathway\\u003c/h3\\u003e\\n\\u003cp\\u003eThe basal culture medium consisted of MS supplemented with 30 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e sucrose and 7 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e agar. Various concentrations of plant growth regulators were added, including 6-benzylaminopurine(6-BA)at 0.5, 1, and 2 mg/l, kinetin (KT) at 0.1, 0.5, and 1 mg/l, and naphthaleneacetic acid (NAA) at 0.01, 0.05, and 1 mg/l. Each treatment involved inoculating 20 leaf samples with three replicates. The differentiation rate was calculated after 30 days culture.\\u003c/p\\u003e\\n\\u003ch3\\u003eCallus Induction\\u003c/h3\\u003e\\n\\u003cp\\u003eThe basal culture medium consisted of MS supplemented with 30 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e sucrose and 7 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e agar. Different combinations of plant growth regulators were added, including 6-BA at concentrations of 0.5, 1, and 2 mg/l, and a combination of 2,4-dichlorophenoxyacetic acid (2,4-D) and indole-3-acetic acid (IAA) at concentrations of 0.1, 0.5, and 1 mg/l each. Each treatment involved inoculating 20 leaf samples, with three replicates. After 30 days, both the callus induction rate and the growth status of calli were assessed.\\u003c/p\\u003e\\n\\u003ch3\\u003eAdventitious shoot differentiation\\u003c/h3\\u003e\\n\\u003cp\\u003eThe basal culture medium comprised MS supplemented with 30 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e sucrose and 7 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e agar. Various combinations of plant growth regulators were added, including 6-BA at concentrations of 1, 2, and 3 mg/l, thidiazuron (TDZ) at concentrations of 0.5, 1, and 2 mg\\u0026middot;\\u003csup\\u003e\\u0026minus;1\\u003c/sup\\u003e, along with combinations of IAA and NAA at concentrations of 0.05, 0.5, and 1 mg/l each. Each treatment involved inoculating 20 pieces of callus tissue, with three replicates. After 30 days, the rates of adventitious shoot and root differentiation, as well as the average number of regenerated shoots, were recorded.\\u003c/p\\u003e\\n\\u003ch3\\u003eAdventitious shoot proliferation\\u003c/h3\\u003e\\n\\u003cp\\u003eThe basal culture medium consisted of MS supplemented with 30 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e sucrose and 7 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e agar. Differentiated adventitious shoots were transferred to media containing combinations of 6-BA at concentrations of 0.5, 1, and 2 mg/l, and NAA at concentrations of 0.01, 0.05, and 0.1 mg/l. Fifteen adventitious shoots were inoculated for each treatment, with three replicates per treatment. After 30 days, the proliferation of adventitious shoots was assessed, and parameters such as proliferation rate, proliferation coefficient, and plant height were measured.\\u003c/p\\u003e \\u003cdiv id=\\\"Sec8\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eAdventitious root formation\\u003c/h2\\u003e \\u003cp\\u003eThe basal culture medium comprised 1/2 MS supplemented with 30 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e sucrose and 7 g\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e agar. The proliferated adventitious shoots were dissected into single shoots, which were then individually transferred to 16 different media containing various concentrations of NAA (0, 0.1, 0.5, 1 mg/l) or IBA (0, 0.1, 0.5, 1 mg/l). Fifteen shoot explants were inoculated for each treatment, with three replicates per treatment. After 30 days, the rooting of shoots was observed, and rooting percentage as well as plant height were recorded.\\u003c/p\\u003e \\u003c/div\\u003e\\n\\u003ch3\\u003eHardening and transplantation\\u003c/h3\\u003e\\n\\u003cp\\u003eAfter removing the tissue-cultured seedlings, approximately 3 cm in height with roots about 2 cm long, they were immersed in distilled water. Following a 2-day placement at room temperature, they were transferred outdoors for a 3-day acclimatization period. Subsequently, they were removed and the culture medium was washed away with distilled water. Finally, they were transplanted into pots containing a 1:1 volume ratio mixture of sterilized vermiculite and nutrient soil.\\u003c/p\\u003e\\n\\u003ch3\\u003eData collection and analysis\\u003c/h3\\u003e\\n\\u003cp\\u003eBrowning rate (%)\\u0026thinsp;=\\u0026thinsp;Number of browning occurrences / Number of inoculations\\u0026times;100%; Callus induction rate(%)\\u0026thinsp;=\\u0026thinsp;Number of explants with callus / Number of inoculations\\u0026times;100%; Differentiation rate(%)\\u0026thinsp;=\\u0026thinsp;Number of calli differentiating into shoots/Number of inoculations\\u0026times; 100%; Shoot proliferation rate (%)\\u0026thinsp;=\\u0026thinsp;Number of shoots proliferating / Number of inoculations\\u0026times; 100%; Average regenerated shoots (per)\\u0026thinsp;=\\u0026thinsp;Total number of regenerated shoots / Total number of regenerated explants; Rooting rate (%)\\u0026thinsp;=\\u0026thinsp;Number of rooted seedlings (or leaves) / Number of surviving seedlings (or leaves) \\u0026times; 100%;\\u003c/p\\u003e \\u003cp\\u003eVariance analysis and significance analysis were conducted using SPSS 23.0 statistical software. Measurement data were calculated and charts were made using Excel 2016. All data represent the average of three repeated experiments.\\u003c/p\\u003e\"},{\"header\":\"Results\",\"content\":\"\\u003cdiv id=\\\"Sec12\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eDirect leaf regeneration\\u003c/h2\\u003e \\u003cp\\u003eThe leaf began to expand on the 7th day of inoculation (Fig.\\u0026nbsp;1A), with two distinct patterns of adventitious root differentiation observed. One emerged subsequent to leaf browning, characterized by dense and short root systems (Fig.\\u0026nbsp;1B) and the other formed prior to leaf browning, exhibiting longer root systems (Fig.\\u0026nbsp;1C).\\u003c/p\\u003e \\u003cp\\u003eThe leaf browning rate ranged from 26.67\\u0026ndash;51.34%, while rooting rates from 13.67\\u0026ndash;72.86%, and budding rates from 0\\u0026ndash;63.46% (Table\\u0026nbsp;\\u003cspan refid=\\\"Tab1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e). Significantly higher rooting rates were observed when 6-BA and KT concentrations were set at 0.5 mg/l, and NAA concentration at 1 mg/l, compared to other treatments (\\u003cem\\u003eP\\u003c/em\\u003e\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05). Notably, leaf browning rates significantly elevated when the KT concentration was higher than the 6-BA concentration (treatment 3) (P\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05)No discernible adventitious bud differentiation but higher rooting rates were observed in treatments 1, 2, and 4. Elevated budding rates (Fig.\\u0026nbsp;1D), lower browning rates, and an average regeneration count of 4.39 were revealed in treatment 7. Collectively, The optimal medium for direct leaf regeneration of adventitious buds was determined to be MS\\u0026thinsp;+\\u0026thinsp;2 mg/l6-BA\\u0026thinsp;+\\u0026thinsp;0.1 mg/lKT\\u0026thinsp;+\\u0026thinsp;0.05 mg/lNAA.\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab1\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 1\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eEffects of various hormone combinations on leaf primordia direct differentiation\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"8\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c7\\\" colnum=\\\"7\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c8\\\" colnum=\\\"8\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eTreatment\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e6-BA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eKT\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eNAA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eBrowning rate(%)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eRooting rate(%)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eGermination rate(%)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAverage regeneration bud count\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.01\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e39.34\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.23\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e55.74\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.33\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e27.14\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.52\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e72.86\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.06\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e51.34\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.54\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e36.76\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.77\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e11.76\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.58\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e1.63\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.26\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e4\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e38.81.\\u0026plusmn;8.59\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e61.19\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.76\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e37.08\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;7.38\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e48.39\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.23\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e16.13\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;3.58\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e1.76\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.35\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e6\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.01\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e40.63\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.22\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e34.38\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;7.98\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e26.56\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.82\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e1.93\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.21\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e7\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e26.67\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.59\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e13.67\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.74\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e63.46\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;12.38\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e4.39\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.14\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e8\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.01\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e29.51\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.71\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e49.18\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.56\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e21.31\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;4.22\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e3.4\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.74\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e9\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e33.87\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;7.89\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e37.10\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.69\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e29.03\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.29\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e2.33\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.42\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003ctfoot\\u003e \\u003ctr\\u003e\\u003ctd colspan=\\\"8\\\"\\u003eNote: Different lowercase letters following data in the same column indicate statistical differences (P\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05).\\u003c/td\\u003e\\u003c/tr\\u003e \\u003ctr\\u003e\\u003ctd colspan=\\\"8\\\"\\u003e\\u0026ldquo;-\\u0026rdquo;: Indicates undifferentiated leaves without adventitious buds\\u003c/td\\u003e\\u003c/tr\\u003e \\u003c/tfoot\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"No\\\" id=\\\"Taba\\\" border=\\\"1\\\"\\u003e \\u003ccolgroup cols=\\\"1\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eFigure\\u0026nbsp;1 The various cultivation stages of direct leaflet differentiation\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003ctfoot\\u003e \\u003ctr\\u003e\\u003ctd colspan=\\\"1\\\"\\u003eNote: A. 7 days post-inoculation; B. Rooting with browning of leaflets observed 30 days post-inoculation; C. Rooting without browning of leaflets observed 30 days post-inoculation; D. Shoot formation observed 30 days post-inoculation; E. Shoot proliferation; F. Rooting initiation on the 8th day; G. Rooting progression on the 15th day; H. Seedling hardening.\\u003c/td\\u003e\\u003c/tr\\u003e \\u003c/tfoot\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec13\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eCallus Induction\\u003c/h2\\u003e \\u003cp\\u003eThe expansion and curling of leaves began on the 7th day after inoculation (Fig.\\u0026nbsp;2A), followed by the induction of callus 15 days later (Fig.\\u0026nbsp;2B). Table\\u0026nbsp;\\u003cspan refid=\\\"Tab2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e reveals that the induction rate of callus from leaves ranged between 19.03% and 86.73% across different hormone combinations in the culture medium. With consistent 6-BA concentrations, the callus induction rate gradually decreased with increasing concentrations of 2,4-D and IAA. At 0.5 and 2 mg/l of 6-BA, the induction rate was higher with the sole addition of 2,4-D compared to IAA, while at 1 mg/l IAA, the induction rate was higher than that with 2,4-D. Among the media containing 6-BA, 2,4-D, and IAA, Treatment 25 exhibited the highest induction rate at 86.73%. The resulting callus appeared yellowish-green and loose. In summary, Treatment 25, with MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.1 mg/l 2,4-D\\u0026thinsp;+\\u0026thinsp;0.5 mg/l IAA, was determined to be the optimal culture medium for inducing callus from leaves.\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab2\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 2\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eEffects of different hormone ratios on leaf callus induction\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"6\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eTreatment\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e6-BA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2,4-D\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eIAA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eInduction rate (%)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eCallus growth state\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e44.17\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.31\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePale yellow, loose, soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e39.98\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.74\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePale yellow, loose, hard texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e19.03\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.33\\u003csup\\u003ed\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow, loose, soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e4\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e49.71\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.63\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e44.54\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.63\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow, compact, hard texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e6\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e38.05\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.94\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePale yellow, loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e7\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e66.11\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.82\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e8\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e53.42\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.44\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePale yellow, loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e9\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e50.76\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;14.13\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePale yellow, loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e10\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e34.52\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.34\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green, loose,soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e11\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e31.96\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.75\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green, compact,softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e12\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e24.07\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;7.22\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green, loose,soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e13\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e51.34\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.99\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green, loose, soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e14\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e46.21\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.23\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green loose softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e15\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e39.14\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.41\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green loose, soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e16\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e55.24\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.91\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green loose, soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e17\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e51.33\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.54\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e18\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e46.61\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;14.26\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e19\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e47.62\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.43\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen, compact, hard texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e20\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e53.41\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.22\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen, compact, hard texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e21\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e55.76\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.16\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen, compact, somewhat hard texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e22\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e42.57\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.25\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e23\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e50.91\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.22\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePale yellow, loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e24\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e44.59\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.17\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green,loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e25\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e86.73\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.29\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow-green,loose, softer texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e26\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e70.23\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;17.05\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eYellow loose, soft texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e27\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e67.15\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.21\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eGreen compact,hard texture\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec14\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eCallus Differentiation of leaf\\u003c/h2\\u003e \\u003cp\\u003eAfter 30 days post-inoculation, callus differentiated into adventitious roots or shoots (Fig.\\u0026nbsp;2C, D). As shown in Table\\u0026nbsp;\\u003cspan refid=\\\"Tab3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e, callus failed to differentiate into shoots when solely supplemented with either 6-BA or TDZ. The rooting rate decreased with increasing concentrations of 6-BA, while it increased with TDZ concentrations, although the differences were not significant. Simultaneous addition of both 6-BA and TDZ yielded shoot differentiation rates ranging from 0\\u0026ndash;7.66%, and rooting rates ranging from 0\\u0026ndash;9.17%. Treatments involving the concurrent addition of 6-BA, TDZ, and NAA resulted in only treatments 16, 17, and 19 differentiating into adventitious roots, while the remaining six treatments exhibited diverse outcomes, with some producing adventitious roots and others generating shoots. Treatment 21 exhibited the highest shoot differentiation rate at 53.48%, while treatment 17 had the highest rooting rate at 23.64%. On average, treatment 21 yielded the highest number of adventitious shoots at 4.23 per explant. Overall, the optimal callus differentiation medium for shoot induction was treatment 21, consisting of MS supplemented with 2 mg/l 6-BA, 2 mg/l TDZ, and 0.5 mg/l IAA.\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab3\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 3\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eEffects of different hormone ratios on leaf callus differentiation\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"7\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"char\\\" char=\\\".\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c7\\\" colnum=\\\"7\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eTreatment\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e6-BA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eTDZ\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eIAA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eGermination\\u003c/p\\u003e \\u003cp\\u003erate(%)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eRooting\\u003c/p\\u003e \\u003cp\\u003erate (%)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAverage number\\u003c/p\\u003e \\u003cp\\u003eof adventitious buds\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e4.58\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.51\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e3.43\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.02\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e2.33\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.93\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.26\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.29c\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e4\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e4.52\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.95\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e6\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e4.74\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.39\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e7\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e1.92\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.82\\u003csup\\u003ed\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.22\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.20\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e8\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e9.17\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.42\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e9\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e6.11\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.21\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e10\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e3.44\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.32\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e8.32\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.31\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.56\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.33\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e11\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e6.41\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.13\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e2.03\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.43\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e12\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e2.91\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.71\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e3.59\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.33\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e13\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e2.37\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.69\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e7.15\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.12\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e3.08\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.93\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e14\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e3.17\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.21\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e15\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e7.66\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.12\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.41\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.22\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e16\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e14.32\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;4.23\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e17\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e23.64\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.34\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e18\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e22.35\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.33\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e4.52\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.96\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e3.27\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.53\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e19\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e17.61\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;3.22\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e20\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e1.32\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.42\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e9.02\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.83\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.93\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.33\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e21\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e53.48\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.89\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e6.49\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.02\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e4.23\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.06\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e22\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e9.46\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.31\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e19.03\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;7.93\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e4.06\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.95\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e23\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e19.83\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.99\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e12.87\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;4.85\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e3.61\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.84\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e24\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e9.10\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2.41\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e8.99\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;3.31\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e2.77\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.32\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec15\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eAdventitious Shoot Proliferation\\u003c/h2\\u003e \\u003cp\\u003eFollowing grafting of adventitious buds onto nine types of bud proliferation culture media, the bud volume increased, and growth rate accelerated. After 20 days, adventitious buds proliferated around the bud blocks. The proliferation rates of adventitious buds via direct differentiation ranged from 7.53\\u0026ndash;87.41%, while those via indirect differentiation ranged from 17.21\\u0026ndash;83.57%(Table\\u0026nbsp;\\u003cspan refid=\\\"Tab4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e). At a concentration of 0.5 mg/l 6-BA, both types of buds exhibited decreased proliferation rates with increasing NAA concentration. Notably, adventitious buds regenerated through the indirect pathway displayed higher proliferation rates. At 1 mg/l, the proliferation rate of adventitious buds via direct differentiation initially increased and then decreased with rising NAA concentration, while the rate via indirect differentiation gradually increased. At 2 mg/l, the proliferation rate of adventitious buds via direct regeneration initially increased and then decreased, whereas the rate via indirect differentiation gradually decreased. The highest proliferation rates for adventitious buds were 87.41% (Fig.\\u0026nbsp;1E) and 82.96% (Fig.\\u0026nbsp;2E), with proliferation coefficients exceeding 4-fold and 3-fold, respectively. Overall, the proliferation rate of adventitious buds via direct regeneration surpassed that via indirect regeneration. Treatment 8, comprising MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.05 mg/l NAA, proved suitable for direct regeneration of adventitious buds, while treatment 7, consisting of MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA and 0.01 mg/l NAA, was deemed suitable for indirect regeneration of adventitious buds.\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab4\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 4\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eEffects of different hormone combinations on adventitious bud proliferation\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"7\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c7\\\" colnum=\\\"7\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colspan=\\\"7\\\" nameend=\\\"c7\\\" namest=\\\"c1\\\"\\u003e \\u003cp\\u003eDirect regeneration Indirect regeneration\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eTreatment\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e6-BA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eNAA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eBud growth\\u003c/p\\u003e \\u003cp\\u003erate (%)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eCoefficient of\\u003c/p\\u003e \\u003cp\\u003eincrement\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eBud growth\\u003c/p\\u003e \\u003cp\\u003erate (%)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eCoefficient of\\u003c/p\\u003e \\u003cp\\u003eincrement\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.01\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e22.53\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;7.95\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e+\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e56.85\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.94\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e16.73\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;3.94\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e+\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e37.16\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;6.77\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e7.53\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1.21\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e+\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e19.81\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;4.31\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e4\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.01\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e69.31\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.97\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e+++\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e65.36\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;12.74\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+++\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e73.86\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.47\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e+++\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e70.39\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.91\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e++\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e6\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e61.91\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.85\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e++\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e76.59\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;14.61\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e++\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e7\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.01\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e81.69\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.61\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e++++\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e82.96\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.85\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+++\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e8\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.05\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e87.41\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;12.56\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e++++\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e74.27\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.81\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+++\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e9\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e82.39\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;15.31\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e++\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e71.51\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;14.33\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e+++\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e++++ means the multiplication factor is more than 4 time +++ means the multiplication factor is more than 3 times; ++ means the multiplication factor is about 2 times; + means the multiplication factor is about 1 times;\\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec16\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eAdventitious Shoot Rooting\\u003c/h2\\u003e \\u003cp\\u003eAfter transferring well-developed seedlings to the rooting medium, root systems formed on the 8th day (Fig.\\u0026nbsp;1F, Fig.\\u0026nbsp;2F), and the roots gradually elongated after 15 days (Fig.\\u0026nbsp;1G, Fig.\\u0026nbsp;2G). As shown in Table\\u0026nbsp;\\u003cspan refid=\\\"Tab5\\\" class=\\\"InternalRef\\\"\\u003e5\\u003c/span\\u003e, among the 10 rooting media, the rooting rate of direct regeneration adventitious buds ranged from 46.17\\u0026ndash;83.69%, while that of indirect regeneration adventitious buds ranged from 43.67\\u0026ndash;79.63%. The rooting rates of adventitious buds for both differentiation pathways increased with higher NAA concentrations and initially increased then decreased with higher IBA concentrations. At the same concentration, the rooting rate with NAA was better than with IBA. When NAA and IBA were combined at both concentrations of 0.1 mg\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e, the highest rooting rates were achieved, 83.69% and 79.63%, respectively, with a decrease in rooting rate as the concentration increased.For the direct regeneration pathway, the plant height of adventitious buds increased with higher NAA and IBA concentrations, while for the indirect regeneration pathway, the plant height first increased then decreased with higher concentrations of NAA and IBA. When NAA and IBA concentrations were both at 0.1 mg\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e, the plant heights of adventitious buds for the two differentiation pathways were 4.52 cm and 3.79 cm, respectively, significantly higher than other treatments (\\u003cem\\u003eP\\u003c/em\\u003e\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05).In summary, NAA has a better effect on adventitious bud rooting of Mimosa pudica than IBA, and the direct regeneration pathway is more effective than the indirect regeneration pathway. The optimal rooting medium for adventitious buds is treatment 8, 1/2 MS\\u0026thinsp;+\\u0026thinsp;0.1 mg\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e NAA\\u0026thinsp;+\\u0026thinsp;0.1 mg\\u0026middot;L\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e IBA. After acclimatization, the plants were transplanted into a substrate mixed with perlite and potting soil in a 1:1 ratio, and the plants grew robustly (Fig.\\u0026nbsp;1H, Fig.\\u0026nbsp;2H).\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab5\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 5\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eEffects of different concentrations of auxins on adventitious bud rooting\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"7\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c7\\\" colnum=\\\"7\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colspan=\\\"7\\\" nameend=\\\"c7\\\" namest=\\\"c1\\\"\\u003e \\u003cp\\u003eDirect regeneration Indirect regeneration\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eTreatment\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eNAA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eIBA\\u003c/p\\u003e \\u003cp\\u003e(mg/l)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eRooting rate(%)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003ePlant height(cm)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eRooting rate(%)\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003ePlant height(cm)\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e56.93\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.51\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e2.63\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.32\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e51.21\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.52\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e2.63\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.32\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e67.39\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;12.79\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e2.86\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.71\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e57.83\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.54\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.62\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.29\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e83.69\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.49\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e3.97\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.85\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e60.36\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.81\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e 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\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e2.42\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.59\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e46.73\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;10.54\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e2.32\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.40\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e73.27\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;14.13\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.44\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.13\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e6\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e59.71\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;12.17\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e2.94\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.77\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e70.26\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.17\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e1.87\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.55\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e7\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e-\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e63.19\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;9.85\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e3.09\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.49\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e51.82\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.62\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e2.53\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.37\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e8\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e74.28\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.19\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e4.03\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.93\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e80.15\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;12.38\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e3.89\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.66\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e9\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e0.5\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e51.66\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.41\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e3.01\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.52\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e71.41\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;11.46\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e3.55\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.46\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e10\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e46.17\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;13.22\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e1.36\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.34\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e51.11\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;8.71\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e2.34\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;0.27\\u003csup\\u003ebc\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"No\\\" id=\\\"Tabb\\\" border=\\\"1\\\"\\u003e \\u003ccolgroup cols=\\\"1\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e\\u003cb\\u003eFigure\\u0026nbsp;2 The various cultivation stages of the indirect leaflet differentiation pathway\\u003c/b\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003ctfoot\\u003e \\u003ctr\\u003e\\u003ctd colspan=\\\"1\\\"\\u003eNote: A. Induction of callus tissue occurs 7 days after inoculation; B. Callus tissue induction is observed 15 days post-inoculation; C. Adventitious roots emerge from the callus tissue 30 days post-inoculation; D. Adventitious shoots develop from the callus tissue 30 days post-inoculation. E. Proliferation of adventitious shoots; F. Rooting begins on the 8th day; G. Rooting progresses by the 15th day; H. Seedling hardening.\\u003c/td\\u003e\\u003c/tr\\u003e \\u003c/tfoot\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e\"},{\"header\":\"Discussion\",\"content\":\"\\u003cp\\u003ePlant regeneration includes two differentiation methods: direct regeneration and indirect regeneration (Long Y et al, 2022). The direct differentiation pathway is the process of selecting explants that do not undergo callus induction and directly differentiate into adventitious buds to obtain regenerated plants; The indirect differentiation pathway involves the induction of callus tissue, adventitious bud differentiation, and rooting of explants to obtain regenerated plants (Jiang W et al, 2012). Direct regeneration path the diameter can obtain regenerated plants in a short period of time, which has the advantages of high efficiency and time saving; The indirect differentiation pathway iswidely used, but the cultivation cycle is long and the workload is large(Youssef N M et al 2019). In \\u003cem\\u003eLamiaceae\\u003c/em\\u003e, indirect regeneration pathways are commonly used, with a cultivation period of over 120 days (Liu J H et al 2022), however, there is relatively little research on direct regeneration. In this study, the regeneration of the indirect differentiation pathway using the leaves of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e pubescens as explants took 120 days, which is similar to the previous research findings. The direct differentiation pathway took only 90 days to obtain regenerated plants, At NAA and IBA concentrations of 0.1 mg/l, the rooting rate and plant height of shoots obtained through direct differentiation pathways are superior to those through indirect pathways. Moreover, compared to the indirect pathway, the direct pathway exhibited stronger genetic stability with no variation in genotype and phenotype traits, making it suitable for regenerating transgenic plants (Hafez et al, 2013 ).\\u003c/p\\u003e \\u003cp\\u003e \\u003cem\\u003eLamiaceae\\u003c/em\\u003e tissue culture commonly uses certain concentrations of cytokinins (6-BA, TDZ, KT)and auxin (NAA, IAA, etc.)2.4-D induced callus and differentiation (Kosar et al 2012, Gharari Z, 2021). The color and quality ofcallus tissue are influenced by both. The impact of concentration, 6-BA and 2, 4-D) combination suitable for \\u003cem\\u003eLavandula angustifolia\\u003c/em\\u003e (Chan Wang et al, 2012). Leaf callus induction, when the ratio of the two is 1:l, results in better callusquality and stronger differentiation ability in the later stage. The concentration of 6-BA and NAA at 10:1 is suitable for inducing callus tissue from \\u003cem\\u003eDracocephalum tanguticum\\u003c/em\\u003e leaves. In previous studies, a ratio of 6-BA to NAA at 1:1 can induce callus formation in the leaves of Phyllostachys sinensis, but its color is yellow and itstexture is loose. In the later stage, only adventitious roots are differentiated, and its ability to further produce adventitious buds is poor (Yue Zong, 2019). In this study, the induction rate and quality of leaf callus tissue showed different states in different hormone combinations. In the combination of 6-BA and asingle combination of 2,4-D or IAA, the induction rate of leaf callus tissue is low, and the callus tissue is in poor condition, which is not conducive to the differentiation of adventitious buds in the later stage. However, in the medium with the addition of 6-BA, 2,4-D and IAA, the induction rate of callus tissue is the highest at 86. 73%. At this time, the callus tissue is loose and the color is yellow green. This is different from previous research results, possibly because the synergistic effect of multiplehormones is much greater than that of a single hormone used alone, and its effect is closely related to the type and concentration of endogenous hormones in the explant andcallus tissue itself (Darvishi et al, 2014; Gharari et al, \\u003cspan citationid=\\\"CR6\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003eDuring the differentiation stage, the mesophyll cells of explants possess dual characteristics of high auxin accumulation and hypersensitivity to cytokinins, enabling the explants to regenerate both roots and shoots. When the medium contains high concentrations of auxin and low levels of cytokinins, the explants differentiate into roots due to the accumulated auxin. Conversely, high concentrations of cytokinins in the medium exploit the hypersensitivity of callus tissues, rapidly and effectively activating shoot gene expression, thereby promoting shoot development (Bidabadi et al 2020). In the direct regeneration of leaves in this study, when NAA concentrations were at 1 mg/l and 6-BA or KT concentrations were at 0.5 mg/l or 0.1 mg/l, a higher rooting rate was observed with no adventitious bud differentiation. Conversely, higher concentrations of 6-BA at 2 mg/l and lower concentrations of NAA at 0.01\\u0026ndash;0.05 mg/l resulted in higher adventitious bud differentiation rates but lower rooting rates. In the indirect regeneration of leaves, higher IAA concentrations at 1 mg/l with 6-BA or TDZ at 1 mg/l led to higher rooting rates but lower adventitious differentiation rates, consistent with the aforementioned research findings. For \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e leaf explants, higher rates of adventitious root differentiation were observed in media containing combinations of 6-BA and NAA, with no adventitious bud formation (Zong Y, 2019), while IAA facilitated bud induction in \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e (Liu J et al, 2017). In this study, the induction rates of adventitious buds in callus tissues were significantly lower with the addition of single or combined 6-BA and TDZ, whereas the addition of IAA markedly enhanced the differentiation rate of adventitious buds, aligning with previous research conclusions (Liu J et al, 2017). This suggests a close relationship between the regeneration of adventitious roots or buds in explants and the types of exogenous hormones used. The high rate of adventitious root differentiation and low rate of adventitious bud differentiation in direct or indirect regeneration processes of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e leaf explants may also be related to the content of endogenous hormones within the plant (Gharari et al, \\u003cspan citationid=\\\"CR6\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e). However, further investigation is required to elucidate the specific changes in endogenous hormone types and levels.\\u003c/p\\u003e \\u003cp\\u003eThe combination of 6-BA and NAA is commonly used in the propagation of \\u003cem\\u003eLabiatae\\u003c/em\\u003e species through adventitious bud culture. Optimal bud proliferation occurs when the concentration of 6-BA ranges from 1 to 2 mg/l and NAA ranges from 0.01 to 0.5 mg/l (Xi Y K et al, 2020 ). At 1 mg/l of 6-BA, the proliferation efficiency of direct mint buds decreases with increasing NAA concentration, whereas at 0.5 mg/l, the efficiency of indirect mint bud proliferation increases with higher NAA concentrations (Islam A et al, 2018). 6-BA plays a crucial role in the proliferation of indirect buds in \\u003cem\\u003eSalvia splendens\\u003c/em\\u003e, but excessive concentrations inhibit bud cluster formation. In media supplemented solely with NAA or IBA, splendens exhibits low rates of adventitious bud proliferation and weaker bud growth (Liu Hui et al, \\u003cspan citationid=\\\"CR14\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e). In this study, at a concentration of 0.5 mg/l of 6-BA, the proliferation rates of both differentiation pathways of adventitious buds increase with higher NAA concentrations. At 1 mg/l, the proliferation of direct pathway buds decreases with increasing NAA, while the indirect pathway shows an opposite trend, consistent with previous findings (Islam A et al, 2018). This indicates that different explant sources treated with the same hormone combination exhibit varying proliferation rates. Selecting appropriate combinations of 6-BA and NAA can yield higher proliferation rates and coefficients (Ghasemiomran et al, 2018).\\u003c/p\\u003e \\u003cp\\u003eAuxin NAA and lBA are commnly used in root culture of plants in the family \\u003cem\\u003eLamiaceae\\u003c/em\\u003e, but the concentration of auxin suitable for plant rooting varies(Aghazadeh et al, 2015; Bassolino L et al, 2015). The rooting effect of IBA at the same concentration on \\u003cem\\u003eElsholtzia bodinieri\\u003c/em\\u003e is better than that of NAA. As the concentration of IBA increases, the rooting rate of \\u003cem\\u003eElsholtzia bodinieri\\u003c/em\\u003e (Zhao Jie et al, \\u003cspan citationid=\\\"CR24\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e) increases with the increase of IBA concentration, The appropriate concentration for its rooting is l mg/l. The rooting effect of NAA at the same concentration on \\u003cem\\u003eSalvia hispanica\\u003c/em\\u003e is better than that of IBA(Li Wei-li et al, 2016). The single addition of NAA and IBA is not conducive to the induction of the root system of \\u003cem\\u003eClerodendranthus spicatus\\u003c/em\\u003e (Li Jia-hui et al, 2022). At the same time, adding l mg/l of NAA and IBA each resulted in a higher rooting rate and robust plant growth, In this study, it was found that the hormone combination added with NAA had a better overall rooting effect than the combination added with IBA. When NAA and IBA were added simultaneously, the rooting rate was higher and the rooting rate of adventitious buds increased with the concentration of NAA and IBA.\\u003c/p\\u003e\"},{\"header\":\"Conclusion\",\"content\":\"\\u003cp\\u003eIn summary, this study established a regeneration system through direct and indirect differentiation pathways using the leaves of tissue cultured seedlings of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e as explants. Research has found that the culture medium for direct regeneration of leaves is MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.1 mg/l KT\\u0026thinsp;+\\u0026thinsp;0.05 mg/l NAA. The differentiation rate of adventitious buds is 63.46%, The callus induction medium is MS\\u0026thinsp;+\\u0026thinsp;2 mg/l6-BA\\u0026thinsp;+\\u0026thinsp;0.1 mg/l 2,4-D\\u0026thinsp;+\\u0026thinsp;0.5 mg/l IAA, with an induction rate of 86.73%. The differentiation medium is MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;2 mg/l TDZ\\u0026thinsp;+\\u0026thinsp;0.05 mg/l IAA, with an adventitious bud differentiation rate of 66.37%.The adventitious bud proliferation medium for the two differentiation pathways is the same. MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.05 mg/l NAA, The proliferation rates for direct differentiation and indirect differentiation pathways were 83.57% and 87.41%, respectively. The optimal rooting medium consisted of 1/2 MS supplemented with 0.1 mg/l NAA and 0.1 mg/l IBA, resulting in rooting rates of 83.69% for direct differentiation and 79.15% for indirect differentiation pathways. The direct differentiation pathway takes 90 days from leaf isolation to plant regeneration, which is 30 days less than the indirect differentiation pathway. The process is simple and the differentiation rate, proliferation rate, and rooting rate are higher than the indirect differentiation pathway. This provides the oretical and technical support for the subsequent research on the genetic transformation system of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e.\\u003c/p\\u003e\"},{\"header\":\"References\",\"content\":\"\\u003col\\u003e\\n\\u003cli\\u003eAghazadeh P, Behboodi B S(2015)Seed germination and in vitro organogenesis optimization of Zhumeria majdae (Lamiaceae) from Iran. American Journal of Plant Sciences, 6(11): 1850-1856.https//:doi: 10.4236/ajps.2015.611185 \\u003c/li\\u003e\\n\\u003cli\\u003eBassolino L, Giacomelli E, Giovanelli S(2015)Tissue culture and aromatic profile in Salvia dolomitica Codd. Plant Cell, Tissue and Organ Culture (PCTOC), 121: 83-95. https//:doi:10.1007/s11240-014-0681-3\\u003c/li\\u003e\\n\\u003cli\\u003eBidabadi S S, Jain S M(2020)Cellular, molecular, and physiological aspects of in vitro plant regeneration. Plants, 9(6): 702. https://doi.org/10.3390/plants9060702\\u003c/li\\u003e\\n\\u003cli\\u003eDeepa A V, Thomas D T(2022). Tissue Culture Studies in Lamiaceae: A Review. Biotechnology and Crop Improvement, 181-211. https://doi:10.1201/9781003239932-11\\u003c/li\\u003e\\n\\u003cli\\u003eDarvishi E, Kazemi E, Kahrizi D(2014)Optimization of callus induction in Pennyroyal (Mentha pulegium). Journal of Applied Biotechnology Reports, 1(3): 97-100. https://doi:10.1023/A:1000304922507.\\u003c/li\\u003e\\n\\u003cli\\u003eGharari Z, Bagheri K, Karimkhanlooei G(2021)Study of tissue culture and in vitro organogenesis of Scutellaria bornmuelleri using benzylaminopurine, lsopentenyl adenine and thidiazuron. South African Journal of Botany, 139(13): 458-469. https://doi.org/10.1016/j.sajb.2021.03.030\\u003c/li\\u003e\\n\\u003cli\\u003eGhasemiomran V, Movahedi M, Alizadeh F(2018)Effects of explant type and plant growth regulator combinations on in vitro direct and indirect regeneration of Stevia rebaudiana.19(23):137-147. https://doi:10.1007/s11627-014-9640-2\\u003c/li\\u003e\\n\\u003cli\\u003eHafez R F, Shahabzadeh Z, Heidari B(2013)Investigation of the efficiency of direct and indirect regeneration in evening primrose (Oenothera biennis). Journal of crop science and biotechnology, 16(09): 291-296. htttps://:doi:10.1007/s12892-013-0115-5\\u003c/li\\u003e\\n\\u003cli\\u003eIslam A, Alam M F(2018)In vitro callus induction and indirect organogenesis of Mentha piperita (L.)-an aromatic medicinal plant. GSC Biological and Pharmaceutical Sciences, 4(3): 049-060. https://doi.org/10.30574/gscbps.2018.4.3.0078\\u003c/li\\u003e\\n\\u003cli\\u003eJiang W, Chen L, Pan Q(2012)An efficient regeneration system via direct and indirect organogenesis for the medicinal plant Dysosma versipellis (Hance) M. Cheng and its potential as a podophyllotoxin source. Acta Physiologiae Plantarum,34(23): 631-639. https://doi:10.1007/s11738-011-0864-z\\u003c/li\\u003e\\n\\u003cli\\u003eKosar M, Mahmoud O(2012)Trichomes and regeneration by direct organogenesis of medicinal plant Dracocephalum kotschyi L. using shoot tips (Lamiaceae). Journal of Crop Science and Biotechnology, 15(3): 251-257. https://doi:10.1007/s12892-012-0019-9\\u003c/li\\u003e\\n\\u003cli\\u003eLong Y, Yang Y, Pan G(2022)New insights into tissue culture plant-regeneration mechanisms. Frontiers in plant science,13(08):752-926. https://doi:10.3389/fpls.2022.926752\\u003c/li\\u003e\\n\\u003cli\\u003eLiu J H, Dong W C, Fei F F( 2022)Regulation of WOX11 expression represents the difference between direct and indirect shoot regeneration. Frontiers in Plant Science, 13: 850726. https://doi.org/10.3389/fpls.2022.850726\\u003c/li\\u003e\\n\\u003cli\\u003eLiu Hui, Chen Yi, Zhao Hang-ping(2011). Elementary study on the high effective regeneration system of Salvia Splendens. Acta Agriculturae Zhejiangensis. 23(02):318-323. https://doi:10.3969/j.issn.1004-1524.2011.02.024.\\u003c/li\\u003e\\n\\u003cli\\u003eLiu Jun, Sun Rui-fen, Geng Mu-dan. et al(2017). Establishment of rapid propagation system of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e Hance in vitro culture. Journal of Northern Agriculture, 45(06): 102-106. https://doi:10.3969/j.issn.2096-1197.2017.06.19.\\u003c/li\\u003e\\n\\u003cli\\u003eLi Wei-li, Lifang Huang, Xinjie Xia(2016)Tissue culture and rapid propagation of Salvia hispanica. Pratacural Science, 33(03):393-399. https:// doi:10.11829/j.issn.1001-0629.2015-0424\\u003c/li\\u003e\\n\\u003cli\\u003eLi Jia-hui, Ye Wei-yan, Zhu Peng-jin, et al(2022)Establishment of a sterile short shoot tissue culture and rapid propagation system for Clerodendranthus spicatus. Chinese Journal of Tropical Crops, 43(10): 2063-2070. http://doi: 10.3969/j.issn.1000-2561.2022.10.012\\u003c/li\\u003e\\n\\u003cli\\u003eTian Xu-ping, Guo Qing-hao, Sun Ming-yu, et al(2021). Comparison on bulbil formation of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e Hance originated from different provenances. Acta Agrestia Sinica, 29(06):1357 - 1362. https://doi:10.11733/j.issn.1007-0435.2021.06.028. \\u003c/li\\u003e\\n\\u003cli\\u003eWang Chan, Chen Li-juan, Cheng Ming-hua, et al(2012). Study on in vitro cultivation of \\u003cem\\u003elavandula angustifolia\\u003c/em\\u003e, Journal of Hainan Normal University（Natural Science）, 25(04): 435\\u0026ndash;437. https://doi:10.3969/j.issn.1674-4942.2012.04.022.\\u003c/li\\u003e\\n\\u003cli\\u003eXi Y K, Wang Y, Zeng B(2020). Callus induction and adventitious bud differentiation of Cyclocodon lancifolius (Roxb.) Kurz. Botanical Sciences, 98(4): 534-544. https://doi:10.17129/botsci.2609\\u003c/li\\u003e\\n\\u003cli\\u003eYoussef N M, Shaaban S A, Ghareeb Z F(2019). In vitro bulb formation of direct and indirect regeneration of Lilium orientalis cv.\\u0026ldquo;Starfighter\\u0026rdquo; plants. Bulletin of the National Research Centre, 43: 1-9.https://doi.org/10.1186/s42269-019-0246-z\\u003c/li\\u003e\\n\\u003cli\\u003eZong Yue, Guo Qing-hao, Tian Xu-ping(2019). Callus Induction and Subculture Test of Stem Segment of \\u003cem\\u003eDracocephalum rupestre.\\u003c/em\\u003e Journal of Shanxi Agricultural Sciences. 47(04):507-509. https:// doi:10.3969/j.issn.1002-2481.2019.04.04.\\u003c/li\\u003e\\n\\u003cli\\u003eZhang Jing, Niu Zhe, Fan Wei-fang (2020). Establishing tissue culture and regeneration system of Scutellaria regeliana. Bulletin of Botanical Research, 40(1): 50-59. https://doi:10.7525/j.issn.1673-5102.2020.01.008\\u003c/li\\u003e\\n\\u003cli\\u003eZhao Jie, Hou Yu-qi, Zhen Wei, Xu Xiao-dan(2023). Study on invitro rapid propagation of Elsholtzia bodinieri. Northern Horticulture, 45(23):60-67. https://doi:10.11937/bfyy.20231147\\u003c/li\\u003e\\n\\u003c/ol\\u003e\"}],\"fulltextSource\":\"\",\"fullText\":\"\",\"funders\":[],\"hasAdminPriorityOnWorkflow\":false,\"hasManuscriptDocX\":true,\"hasOptedInToPreprint\":true,\"hasPassedJournalQc\":\"\",\"hasAnyPriority\":false,\"hideJournal\":false,\"highlight\":\"\",\"institution\":\"\",\"isAcceptedByJournal\":true,\"isAuthorSuppliedPdf\":false,\"isDeskRejected\":\"\",\"isHiddenFromSearch\":false,\"isInQc\":false,\"isInWorkflow\":false,\"isPdf\":false,\"isPdfUpToDate\":true,\"isWithdrawnOrRetracted\":false,\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"plant-cell-tissue-and-organ-culture-pctoc\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":false,\"externalIdentity\":\"pcto\",\"sideBox\":\"Learn more about [Plant Cell, Tissue and Organ Culture (PCTOC)](https://www.springer.com/journal/11240)\",\"snPcode\":\"11240\",\"submissionUrl\":\"https://submission.nature.com/new-submission/11240/3\",\"title\":\"Plant Cell, Tissue and Organ Culture (PCTOC)\",\"twitterHandle\":\"\",\"acdcEnabled\":true,\"dfaEnabled\":true,\"editorialSystem\":\"em\",\"reportingPortfolio\":\"Springer Hybrid\",\"inReviewEnabled\":true,\"inReviewRevisionsEnabled\":false},\"keywords\":\"Dracocephalum rupestre, differentiation pathways, callus induction, proliferation, rooting\",\"lastPublishedDoi\":\"10.21203/rs.3.rs-4968870/v1\",\"lastPublishedDoiUrl\":\"https://doi.org/10.21203/rs.3.rs-4968870/v1\",\"license\":{\"name\":\"CC BY 4.0\",\"url\":\"https://creativecommons.org/licenses/by/4.0/\"},\"manuscriptAbstract\":\"\\u003cp\\u003eLeaf explants of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e were utilized for regeneration employing direct and indirect differentiation pathways. Results revealed that the direct regeneration medium for leaf explants consisted of MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.1 mg/l KT\\u0026thinsp;+\\u0026thinsp;0.05 mg/l NAA, yielding a differentiation rate of 63.46%. The induction medium for callus was composed of MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.1 mg/l 2,4-D\\u0026thinsp;+\\u0026thinsp;0.5 mg/l IAA, resulting in an induction rate of 86.73%. For the differentiation of adventitious buds, the medium included MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;2 mg/l TDZ\\u0026thinsp;+\\u0026thinsp;0.05 mg/l IAA, with a differentiation rate of 53.48%. The proliferation medium for adventitious buds generated through both pathways, comprised MS\\u0026thinsp;+\\u0026thinsp;2 mg/l 6-BA\\u0026thinsp;+\\u0026thinsp;0.05 mg/l NAA, with proliferation rates of 83.57% and 87.41%, respectively. The rooting medium suitable for both methods was 1/2MS\\u0026thinsp;+\\u0026thinsp;0.1 mg/l NAA\\u0026thinsp;+\\u0026thinsp;0.1 mg/l IBA, resulting in rooting rates of 83.69% and 79.15%, respectively. Comparatively, the direct differentiation pathway proved to be more efficient and time-saving, with leaf explants requiring 30 days less for regeneration compared with the indirect pathway. This study provides theoretical and technical support for subsequent genetic transformation research of \\u003cem\\u003eDracocephalum rupestre\\u003c/em\\u003e.\\u003c/p\\u003e\",\"manuscriptTitle\":\"Regeneration of in vitro plants through direct and indirect organogenesis from Dracocephalum rupestre leaf explants\",\"msid\":\"\",\"msnumber\":\"\",\"nonDraftVersions\":[{\"code\":1,\"date\":\"2024-12-02 12:54:47\",\"doi\":\"10.21203/rs.3.rs-4968870/v1\",\"editorialEvents\":[{\"type\":\"communityComments\",\"content\":0},{\"type\":\"reviewerAgreed\",\"content\":\"\",\"date\":\"2024-11-15T09:27:32+00:00\",\"index\":0,\"fulltext\":\"\"},{\"type\":\"reviewersInvited\",\"content\":\"\",\"date\":\"2024-11-14T12:10:21+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorAssigned\",\"content\":\"\",\"date\":\"2024-10-21T14:59:44+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"submitted\",\"content\":\"Plant Cell, Tissue and Organ Culture (PCTOC)\",\"date\":\"2024-10-21T05:46:51+00:00\",\"index\":\"\",\"fulltext\":\"\"}],\"status\":\"published\",\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"plant-cell-tissue-and-organ-culture-pctoc\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":false,\"externalIdentity\":\"pcto\",\"sideBox\":\"Learn more about [Plant Cell, Tissue and Organ Culture (PCTOC)](https://www.springer.com/journal/11240)\",\"snPcode\":\"11240\",\"submissionUrl\":\"https://submission.nature.com/new-submission/11240/3\",\"title\":\"Plant Cell, Tissue and Organ Culture (PCTOC)\",\"twitterHandle\":\"\",\"acdcEnabled\":true,\"dfaEnabled\":true,\"editorialSystem\":\"em\",\"reportingPortfolio\":\"Springer Hybrid\",\"inReviewEnabled\":true,\"inReviewRevisionsEnabled\":false}}],\"origin\":\"\",\"ownerIdentity\":\"0389a31c-99a3-4801-b13b-394f42a61046\",\"owner\":[],\"postedDate\":\"December 2nd, 2024\",\"published\":true,\"recentEditorialEvents\":[],\"rejectedJournal\":[],\"revision\":\"\",\"amendment\":\"\",\"status\":\"published-in-journal\",\"subjectAreas\":[],\"tags\":[],\"updatedAt\":\"2025-03-31T16:06:28+00:00\",\"versionOfRecord\":{\"articleIdentity\":\"rs-4968870\",\"link\":\"https://doi.org/10.1007/s11240-025-03026-1\",\"journal\":{\"identity\":\"plant-cell-tissue-and-organ-culture-pctoc\",\"isVorOnly\":false,\"title\":\"Plant Cell, Tissue and Organ Culture (PCTOC)\"},\"publishedOn\":\"2025-03-25 15:57:10\",\"publishedOnDateReadable\":\"March 25th, 2025\"},\"versionCreatedAt\":\"2024-12-02 12:54:47\",\"video\":\"\",\"vorDoi\":\"10.1007/s11240-025-03026-1\",\"vorDoiUrl\":\"https://doi.org/10.1007/s11240-025-03026-1\",\"workflowStages\":[]},\"version\":\"v1\",\"identity\":\"rs-4968870\",\"journalConfig\":\"researchsquare\"},\"__N_SSP\":true},\"page\":\"/article/[identity]/[[...version]]\",\"query\":{\"redirect\":\"/article/rs-4968870\",\"identity\":\"rs-4968870\",\"version\":[\"v1\"]},\"buildId\":\"8U1c8b4HqxoKbykW_rLl7\",\"isFallback\":false,\"isExperimentalCompile\":false,\"dynamicIds\":[84888],\"gssp\":true,\"scriptLoader\":[]}","source_license":"CC-BY-4.0","license_restricted":false}