Study on the Anti-Aging Physiological Characteristics and Molecular Mechanism of Camellia Oleifera

Study on the Anti-Aging Physiological Characteristics and Molecular Mechanism of Camellia Oleifera | 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 Study on the Anti-Aging Physiological Characteristics and Molecular Mechanism of Camellia Oleifera Yongzhong Chen, Jianjun Chen, Zhen Zhang, Yanming Xu, Zhilong He, and 9 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-493148/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract To study the physiological and molecular regulating mechanism of ancient Camellia oleifera which kept a exuberant vitality for more than one hundred years, leaves of 30a year old and > 100 year old Camellia oleifera were selected as targets. On the basis of the study of the generation and the clearance of reactive oxygen species, sequencing analysis of the transcriptome and expression profiling by high throughput sequencing analysis technique was conducted to study differentially expressed functional genes related to the tree age. It showed that the chlorophyll content and enzyme activities increased in ancient Camellia oleifera leaves. Expression of chlorophyll a/b binding protein gene, auxin related gene, the signal transduction factor and the transcription factor gene in ancient trees were all higher than mature tree. The down regulated gene expression of inductive genes related to protein degradation in ancient tree. Under the comprehensive function of those factors, ancient Camellia oleifera leaves still kept an exuberant vitality which was very useful for studies of stress resistance molecular biology and genetic improvement of Camellia oleifera. Plant Molecular Biology and Genetics Camellia oleifera transduction factor exuberant vitality sequencing analysis technique Figures Figure 1 Figure 2 Introduction Tree senescence refers to the process of decline occurring in plants or some organs during growth and development, which leads to the natural termination of life activities 1 , 2 . Usually, during tree growth and development, the changes of internal factors, such as hormones, metabolic levels, aging-related genes, transcription factors and the effects of external environment factors, such as drought, high temperature, pests may lead to chlorophyll degradation, protein, lipids, nucleic acid, starch and other biological macromolecules hydrolysis 3 , 4 , and cause, accelerate aging, and even cause tree death. At present, the research on the mechanism of tree senescence includes stomatal regulation theory, senescence gene regulation theory, active oxidation theory, nutrition deficit theory, plant hormone regulation theory, external stress and pest theory 5 , 6 . The commonness of these theories is to acknowledge that plant senescence and death are the universal laws of life development, and are an active and necessary process in plant growth and development, morphogenesis and environmental response. Moreover, these theories also show that tree senescence is formed under the combined action of many different factors. At the same time, studies have found that different tree senescence processes are different due to different mechanisms and inducing factors of senescence 7 . When encountering many external stress and internal senescence related factors, some plants could form the corresponding resistance as well as the defense mechanism by some antioxidase, the transcription factor, and the signal transduction factor and thus prolonged its senescence process. Camellia oleifera which were plants of Camellia family, Camellia plants and perennial small trees or shrubs was a specific oleiferous tree species of China which usually distributed at low mountains and hills in South of China 8 . Elaeis guineensis , Olea europaea , Cocos nucleifera as well as Camellia oleifera were four edible oleiferous tree species of the world. Camellia oleifera possessed characteristics of poor soil tolerance, strong resistance, and exuberant vitality. It was common to find the growth of Camellia oleifera with more than 100 year old age in natural forests. For example, there was a Camellia oleifera tree with more than 120 year old age grown in Hendong Country, Henyang City in China (N27 o 05’, E112 o 56’). This single tree produced annual average Camellia fresh fruit for more than 200 kg during the continuous field observation from 2014 to 2016. What were the factors which maintained the exuberant vitality of those ancient Camellia oleifera ? Nowadays, studies about the transcription level of plants antisenescence mainly focused on the model plants such as Arabidopsis thaliana 9 , 10 . However, research about antisenescence of Camellia oleifera was scarce. In this study, focusing on antisenescence of Camellia oleifera , leaves of Camellia oleifera with different ages were selected as targets. ROS and antioxidase activities in leaves of mature tree (30 a) and ancient tree (> 100 a) were determined. By conducting the transcriptome sequencing analysis, the ROS clearance system of Camellia oleifera with different ages as well as the expression of differential genes was studied. Furthermore, antisenescence physiological characteristics and molecular mechanism of ancient Camellia oleifera were studied preparing for the further studies of stress resistance molecular biology and genetic improvement. Materials And Methods Experimental materials The experimental site was located in the experimental forest farm of Hunan Academy of Forestry (east longitude 113 o 01 , 20 ,, , north latitude 28 o 06 , 40 ,, ). On April 18th, 2017, good single plant of Camellia oleifera of mature tree (30 a) and ancient tree (> 100 a) with strong growth and good development were sampled (Fig. 1). Annual leaves which received consistent illumination, were strong and had no pests and diseases in those single plants at the middle and upper part of the periphery of crown were selected as experimental material, which had three biological replicates. The collected leaves were put into the plastic bags, numbered, quickly put into the ice box and taken back to the laboratory. After rinsing by deionized water and treatment with liquid nitrogen, leaves were put into − 70 o C refrigerator for further analysis. Determination methods Contents of chlorophyll (Chl), malondialdehyde (MDA), hydrogen peroxide (H 2 O 2 ), SOD, POD and CAT were determined by the extraction of acetone, the thiobarbituric acid method, the trichloroacetic acid colorimetry, the riboflavin-NBT photoreduction, the guaiacol method and the permanganate titration method, respectively. Samples were sent to Beijing Genomics Institution in Guangzhou to conduct the transcriptome sequencing. The paired end sequencer of Illumina Hiseq TM 2000 was used for sequencing. Raw data were processed by Beijing Genomics Institution in Guangzhou. Data analysis Excel 2003 was used for the data processing and the figure drawing. SPSS 16.0 was used for data statistics and the variance analysis. The differential significance of the physiological and biochemical indexes was analyzed by one-way ANOVA test. Multiple comparisons used the least significant difference test (LSD). Statement We confirm that the use of plants in the present study complies with international, national and/or institutional guidelines. Results Antisenescence physiological indexes analysis of Camellia oleifera trees with different ages There was a significant difference of chlorophyll content among Camellia oleifera leaves with different ages. Chl a + Chl b content in leaves of ancient Camellia oleifera was 0.44 mg/g, which was higher than 0.27 mg/g in mature tree. Ratio of Chl a to Chl b in leaves of ancient Camellia oleifera were higher than mature tree, which were 10.32、6.34, respectively. MDA and H 2 O 2 contents in leaves of ancient Camellia oleifera were also higher than mature tree, which were 8.83 nmol/g and 126.77 ug/g. Antioxidase activities of SOD, POD and CAT were the highest in ancient Camellia oleifera leaves, namely 671.00 (U/g), 22.58 U/(g min) and 498.42 U/(g min), respectively (Table 1). CAT was specific-functional enzyme for the clearance of H 2 O 2 . POD was also very important for the clearance of surplus H 2 O 2 in plants. The changing trends of these two enzymes were negatively correlated with the change of H 2 O 2 to some extent, illustrating that these two could promote the defense of Camellia oleifera against the attack of H 2 O 2 and thus prolong senescence. SOD mainly cleared superoxide anion radical (O 2 - ). In all, with the increase of the tree age, MDA and H 2 O 2 contents of Camellia oleifera leaves increased, which resulted in excessive or unbalanced ROS in plants and final senescence. Meantime, chlorophyll in ancient Camellia oleifera leaves maintained at a relatively higher level. Besides, ancient Camellia oleifera leaves possessed relatively higher antioxidase activities, which not only effectively guaranteed photosynthesis but also defended the possibly adverse effect caused by the increase of MDA and H 2 O 2 contents. Finally, the normal growth and development was promoted and the senescence process was prolonged. Table 1 Chlorophyll content in Camellia oleifera leaves of different tree ages Chl a/Chl b Chl a+ Chl b (mg/g) MDA (nmol/g) H 2 O 2 (μg/g) SOD (U/g) POD(U/(g·min)) CAT(ug/(g·min)) Ancient tree 10.32a(0.44) 0.44a (0.003) 8.83a (1.25) 126.77a (4.75) 671.00a (35.82) 22.58a (1.01) 498.42a (34.45) Mature tree 6.34c(0.20) 0.27c (0.007) 6.51ab (0.63) 109.43b (1.99) 519.94a (17.56) 14.34ab (1.00) 363.80ab (19.40) Note: Different letters illustrated the difference of physiological and biochemical indexes among different tree ages (P<0.05). Data in brackets were standard deviation. The differential expression of antisenescence related genes of Camellia oleifera leaves in stages of different tree ages The differential expression of chlorophyll degradation related genes Chlorophyll a/b binding protein gene was very important for the maintenance of the chlorophyll content and the photosynthetic ability of the plant. In this study, 10 differentially expressed related genes of chlorophyll a/b-binding protein were found in ancient Camellia oleifera leaves, wherein 9 genes were up regulated expressed and 1 gene was down regulated expressed (Table 2). The expression increase of Chlorophyll a/b binding protein gene possibly prolonged the senescence process by enhancing the photosynthetic ability of ancient Camellia oleifera tree. Table 2 Differential expression of chlorophyll a/b-binding protein related genes Gene category Main functions Differential gene Up regulated gene amount(Ancient tree VS Mature tree) Down regulated gene amount(Ancient tree VS Mature tree) Chlorophyll a/b-binding protein Maintain chlorophyll content and photosynthetic capacity of plants 10 9 1 The differential expression of antioxidase related genes SOD, POD, CAT and APX were common antioxidases, which were usually very important for the clearance of surplus ROS in plants and the prolongation of senescence. In this study, 22 antioxidase genes were found significantly differentially expressed in ancient Camellia oleifera leaves, wherein 15 antioxidase genes were up regulated expressed and 7 genes were down regulated expressed (Table 3). These antioxidase genes mainly included SOD, POD, CAT and APX. POD had the most differential genes amount, followed by those of APX, SOD and CAT. The increase of the expression of antioxidase genes could prolong the senescence process by enhancing the antioxidase activity of ancient Camellia oleifera trees. Table 3 Differential expression of antioxidase related genes Gene categories Main functions Differential gene amount Up regulated gene amount(Ancient tree VS Mature tree) Down regulated gene amount(Ancient tree VS Mature tree) SOD To clear surplus radicals 2 0 2 POD To clear ROS and enhance the stress resistance to drought etc. 10 9 1 CAT To clear surplus H 2 O 2 1 0 1 APX To clear ROS 9 6 3 Total 22 15 7 The differential expression of hormone related genes Hormone levels were closely related to plant senescence. Usually, Auxin and cytokinin (CTK) could prolong plant senescence, whereas abscisic acid (ABA) could accelerate plant senescence. In this study, dramatic differential expression of 40 hormone related genes were found in ancient Camellia oleifera leaves, wherein 21 genes were up regulated and 19 genes were down regulated expressed (Table 4). From categories, hormone related genes mainly included ABA, auxin, CKX and SAUR, wherein auxin related differential gene number was the highest, which was followed by that of ABA. Due to the combined effects of those hormone related genes, ancient Camellia oleifera could maintain hormones at a relatively stable level and further its exuberant vitality. Table 4 Differential expression of hormone related genes Gene categories Main functions Differential gene amount Up regulated gene amount(Ancient tree VS Mature tree) Down regulated gene amount(Ancient tree VS Mature tree) ABA Inhibition of plant growth, promotion of leaf abscission and acceleration plant dormancy 12 6 6 Auxin Promotion of the generation of lateral and adventitious root adjustment of flowering and sex differentiation, adjustment of fruiting and fruit development, and control of apical dominance 25 13 12 CKX Maintenance or reestablishment of the stability and equilibrium of CTK in plants, enhancement of antioxidation ability of plants 1 1 0 SAUR Maintenance of auxin level, adjustment of auxin transport and cell amplification 2 1 1 Total 40 21 19 The differential expression of stress resistance related genes The transcription factor, the signal transduction factor, the resistance gene and the defense related gene could significantly enhance the resistance of plants and further effectively answer the external adverse growth factors, respectively. 496 stress resistance related genes were significantly differentially expressed in ancient Camellia oleifera leaves, wherein 301 genes were up regulated expressed and 195 genes were down regulated expressed. Those stress resistance genes mainly included transcription factors of MYB, NAC, MRKY and zinc Finger etc., signal transduction factors of GTP-binding, receptor protein kinase etc., resistance protein of pathogenesis related protein, disease resistance protein etc., and defense-related gene. The amount of differential genes related to Zinc Finger and disease resistance protein was the highest, followed by other genes of LRR receptor-like serine/threonine-proteinkinase, HSP, MYB, Leucine zipper, bHLH and ERF (Table 5). With the combined effects of those stress resistance related genes, ancient Camellia oleifera could maintain normal growth through the corresponding adjustment and adaption mechanism to response the external growth stress. Table 5 The differential expression of stress resistance related genes Gene categories Main functions Differential gene amount Up regulated gene amount(Ancient tree VS Mature tree) Down regulated gene amount(Ancient tree VS Mature tree) Zinc Finger Defense against drought, high temperature, salt stress, pathogen 128 73 55 HSP Defense against high temperature and drought 39 24 15 MYB Defense against drought, salt, coldness, high temperature stress and enhancement of POD activities of plants 30 15 15 bHLH Clearance of ROS and further enhancement of coldness resistance of plants 31 22 9 Leucine zipper Defense against drought and salt stress 21 18 3 WRKY Defense against drought and salt stress 20 18 2 NAC Defense against drought, high temperature, pathogenic bacteria 10 10 0 MADS-box Defense against drought and water stress 4 3 1 LRR receptor-like Serine/threonine-proteinkinase Defense against drought and salt stress 74 41 33 ERF Enhancement of disease resistance of plants 21 10 11 GTP-binding protein Enhancement of stress resistance and disease resistance of plants 19 6 13 Receptor protein kinase Defense against drought and salt marsh stress 8 5 3 Disease resistance protein Enhancement of disease resistance 56 36 20 Pathogenesis related protein Defense against various pathogenic bacteria 13 7 6 Chitinase Enhancement of disease resistance of plants 8 6 2 Stress protein Enhancement of resistance of plants to external stress 14 7 7 Total 496 301 195 The differential expression of protein degradation related genes Protein degradation which was also an important reason for plant senescence usually occurred due to the function of some proteinase and F-box family protein genes. In this study, 72 functional genes related to protein degradation were found in ancient Camellia oleifera leaves (Table 6). In detail, those included 17 aspartyl protease genes, 11 cysteine proteinase genes and 44 F-box family protein genes. Among cysteine proteinase, 7 genes were up regulated and 4 were down regulated. Among F-box family protein genes and aspartyl protease genes, the expression of most differential genes demonstrated a down regulated trend. With the combined effects, protein degradation of ancient Camellia oleifera leaves may be effectively delayed, which further was possibly helpful for the prolongation of senescence. Table 6 Differential expression of protein degradation related genes Gene categories Main functions Differential gene amount Up regulated gene amount(Ancient tree VS Mature tree) Down regulated gene amount(Ancient tree VS Mature tree) F-box Involvement in protein degradation of plant cells 44 17 27 Aspartyl protease Involvement in senescence and pathogen related protein degradation 17 6 11 Cysteine proteinase Involvement in protein hydrolysis 11 7 4 Total 72 30 42 Verification of transcriptome differential genes Real time quantitative PCR was used to further verify the reliability of transcriptome sequencing results and the expression patterns of differential genes. The selected verification genes include NAC, F-box, aspertyl protection, cystaine protonase, chlorophyll a / B and others. ETIF3H was selected as the internal reference gene. The results showed that the expression of c184012_g3 (F-box)、c178940_g2 (Aspartyl protease)、c163401_g1 (Cysteine proteinase) were down regulated in ancient trees, up regulated in mature tree, c167863_g1 (NAC)、c184170_g1 (Chlorophyll a/b) were up regulated in ancient trees and down regulated in mature tree (Figure 2). In general, the expression patterns of these genes are consistent with the results of transcriptome sequencing, which further verifies the reliability of transcriptome sequencing results. Discussion The Chlorophyll content of Camellia oleifera leaves in ancient trees was significantly higher than those in mature tree. 9 chlorophyll a/b binding proteins in ancient Camellia oleifera leaves demonstrated an up regulated trend. Chlorophyll guaranteed high efficient photosynthesis of plants. Thus, the degradation of chlorophyll was also a main marker of senescence of plant leaves. In Zea mays 11 , Sorghum bicolor 12 , and Arabidopsis thaliana 13 , chlorophyll contents were closely related to leaf senescence. Besides, during leaf senescence, some genes involved in photosynthesis would be down regulated expressed, resulting in the decrease of the photosynthetic ability of leaves 14 , 15 . Chlorophyll binding protein genes were in a down regulated trend during the senescence process of Oryza Sativa 16 and Gossypium 17 . In this study, the chlorophyll content and the expression of chlorophyll a/b binding protein gene of ancient Camellia oleifera leaves demonstrated an increasing trend, illustrating that ancient Camellia oleifera leaves still had a high level of chlorophyll, which was very important for the maintenance of the photosynthesis of the whole tree, the promotion of the growth and development of tree and the prolongation of senescence. Antioxidase activities of SOD, POD and CAT as well as the expression of POD、APX antioxidase related genes in ancient Camellia oleifera leaves demonstrated an increasing trend. In the senescence process of Triticum aestivum 18 and Oryza sativa 19 , activities of SOD, CAT and APX decreased, which will lead to the increase of ROS level and accelerate leaf senescence. A certain concentration of ROS was necessary for the normal physiological process of plants, whereas the excessive ROS would influence and accelerate the senescence process of plants. Thus it was very important to maintain the expression equilibrium of ROS in plants for not only the growth and development but also the prolongation of senescence. Antioxidase and related functional gene expression could effectively clear ROS generated during senescence in leaves 18 , 20 . In this study, multiple antioxidase activities as well as the expression of antioxidase related gene increased in ancient Camellia oleifera leaves. Surplus ROS in ancient Camellia oleifera leaves could be cleared in time, which could decrease the cell harm of ancient Camellia oleifera tree from surplus ROS, further maintain normal physiological metabolism of Camellia oleifera and prolong the senescence of plants. Hormones were very important for the regulation of the growth and development of plants as well as the senescence process of leaves. ABA gene was up regulated expressed in the senescence process of Gossypium 17 , 21 , which could influence the senescence of plants by the regulation of growth stress and the induction of ethylene expression 22 , 23 . Abscisic acid 8’-hydroxylase which was a key functional enzyme, usually was negatively correlated with ABA content in plants 24 . The increase of CTK level in plants can further prolong and inhibit leaf senescence by ways of decreasing Rubisco content as well as enhancing heat resistance, drought resistance, antioxidation ability and photosynthetic ability of plants 24 , 25 , 26 , 27 , 28 . In this study, the expression of 1 CKX related gene increased in ancient Camellia oleifera leaves. In Nicotiana tabacum 29 , the increase of the expression of AtCKX could effectively enhance the antioxidation ability of plants. Auxin related genes could prolong plant senescence by inducing auxin contents in plants and adjusting the NAC transcription factors amount 30 , 31 . In Gossypium , the increase of the expression of auxin related gene could inhibit leaf senescence 17 , 32 . In this study, the expression of 1 SAUR related gene increased in ancient Camellia oleifera leaves. SAUR gene played an important role in the maintenance of auxin level, the regulation of auxin transport and cell amplification 33 , 34 . With the combined effects of those hormone genes, hormone levels in Camellia oleifera leaves could be maintained in balance, which could promote the benign growth and prolong senescence. External growth stresses of high temperature, low temperature, drought, diseases and pests were usually closely related to plant senescence 35 . Due to many external growth stresses, the transcription factors related to stress in plants would be induced to be expressed. The stress transcription factors could regulate the expression of the stress resistance gene in plants, thus enhancing the stress resistance of plants. In Arabisopsis thaliana 36 , Gossypium 17 , Oryza sativa 37 , the increase of the expression of the NAC transcription factor was very useful for prolonging plant senescence, and answering drought as well as salt and marsh stress. The MYB transcription factor expression could enhance the resistance to growth stress of drought, salt, coldness and high temperature in Lycopersicon esculentum 38 , Arabidopsis thaliana 39 , 40 , 41 and Salicornia brachiata 42 . In Oryza sativa 43 and Arabidopsis thaliana 44 , it has been found that WRKY could influence leaf senescence by enhancing the resistance to the adverse factors of growth including drought, high temperature, pathogenic bacteria etc. In Trifoliate Orange , the increase of the expression of the bHLH transcription factor could enhance POD activity of plants, clear ROS, and further enhance coldness resistance of plants 45 . In Brassica rapa 46 .and Lycopersicon esculentum 47 ., it has been found that the increase of the expression of MADS-box gene could enhance the resistance of plants to drought and salt stress. The excessive expression of the HSPs transcription factor was helpful for the enhancement of the stress resistance of plants to high temperature and drought 48 , 49 , 50 .. In Arabidopsis thaliana 51 . and Populous Trichocarpa 52 ., it has been found that the increase of the expression of the functional gene of Zinc Finger protein could enhance the stress resistance of plants to drought, high temperature, salt stress and pathogen by the adjustment of antioxidase activities of plants 53 , 54 .. Zhu et al. found that the slbzip1 transcription factor of Leucine zipper could play an important role in the resistance of Lycopersicon esculentum to salt and drought by regulating the pathway mediated by ABA 55 .. Similar with the transcription factor, plant received stress stimulation, leaded to the physiological change of plant cells through many signal transduction pathways and finally adapted to the change of external environment 56 , 57 .. GTP binding protein was very important for the protection of cell signal transduction and the enhancement of the stress resistance and the disease resistance of plants 58 , 59 . It showed that receptor protein kinase played an important role in growth stress resistance to drought and salt marsh as well as stress responses of antioxidation defense and pathogen reaction 57 , 60 , 61 .. LRR receptor-like serine/threonine proteinkinase (FLS2) usually played an important role in the interaction pathway of plant-pathogen, involved in the defense system of pathogen and finally greatly enhanced the disease resistance ability of plants 62 .. Ethylene-responsive transcription factors (ERF) not only involved in the plant sescenence and the regulation of many stress response but also took part in the defense and stress answering reaction of plants 63 , 64 , 65 .. In the growth process, plant would not only be influenced by external growth stress but also encountered the invasion of many pathogenic bacteria, which finally exacerbated the senescence process. Chitinase was verified to effectively inhibit activities of some pathogens, intensify the defense system of plants and enhance the disease resistance of plants to many pathogenic bacteria 66 , 67 . when encountering stress, plant cells could also increase the defense gene expression of resistance protein, defense-related gene etc. to prevent cells from harm. In ancient trees, the whole gene expression of disease resistance protein and pathogenesis related protein demonstrated an increasing trend. In this study, the expression of multiple signal transduction factors, transcription factors and disease resistance related genes were up regulated. With the combined effect of those functional genes, Camellia oleifera could response to external stress and some pathogen invasion in time, enhance resistance to various growth stresses and pathogenic bacteria through those factors and finally prolong senescence. There were many differential genes related to stress resistance of Camellia oleifera , which was related to the growing environment of Camellia oleifera itself. Camellia oleifera was a specific economic tree species in China, which mainly distributed at low mountains and hills in different provinces of South China. In those areas, during the growth process, Camellia oleifera encountered stresses of low temperature in winter, lots of rain in spring, high temperature in summer and drought. Besides, Camellia oleifera was susceptible to anthracnose and soft rot. By preliminary speculation, due to the influence of those growth stresses and pathogenic bacteria, the expression of many resistance related genes in Camellia oleifera including the transcription factors, the resistance factors, the defense factors and the signal transduction genes, which could better defend against those adverse growth factors and finally promote normal growth and development of plants. In ancient Camellia oleifera leaves, the overall expression of functional genes related to protein degradation were down regulated. Similar with the situation of chlorophyll degradation, protein would also be degraded during plant senescence 15 . Cysteine proteinase was regarded as the most abundant enzyme which was related to plant leaf senescence 68 . and in protein hydrolysis 69 . The expression of aspartic protease which took part in petal senescence as well as pathogen related protein degradation 69 would be increased during senescence in Glycine max 69 and Arabidopsis thaliana 70 . F-box gene was one of the largest gene families which took part in cell protein degradation in plants. Encountering stresses and during leaf senescence, F-box gene was up regulated expressed in chickpea 71 and Oryza sativa 72 . With the degradation of protein, the senescence process of plants would be exacerbated. In ancient Camellia oleifera leaves, the expression of aspartyl protease gene, cysteine proteinase gene and F-box family protein gene were decreased, which effectively inhibited the degradation of protein and thus prolonged senescence. Conclusions This study shows that: (1) With the increase of the tree age of Camellia oleifera , the chlorophyll content in ancient Camellia oleifera leaves was significantly higher than those in mature and young trees, which was also verified by the chlorophyll related gene expression in Camellia oleifera leaves. Multiple chlorophyll a/b-binding proteins in ancient Camellia oleifera leaves demonstrated an up regulated trend, illustrating more active vital movement in ancient Camellia oleifera leaves. (2) The whole expression of related genes of POD, CAT and APX in ancient Camellia oleifera leaves demonstrated an up regulated trend, which was verified by antioxidase contents of SOD, POD and CAT in ancient Camellia oleifera leaves, illustrating that the clearance ability of ROS in ancient Camellia oleifera leaves could be enhanced by the increase of antioxidase activities. (3) The expression of multiple antisenescene hormone related genes increased in Camellia oleifera leaves, wherein auxin related gene number was the most. (4) Multiple stress resistance and pathogenic bacteria resistance related genes were found in ancient Camellia oleifera leaves. With the combined effects of those genes, Camellia oleifera could answer various external growth stress and pathogenic bacteria in time and further response correspondingly by the corresponding transcription factor, signal transduction, resistance protein and defense related gene. (5) The whole expression of the functional differential genes of Aspartyl protease, Cysteine proteinase and F-box family protein, which influenced protein degradation, demonstrated a decreasing trend in ancient Camellia oleifera leaves. Thus, protein degradation would be effectively inhibited. (6) The results of real-time quantitative PCR were consistent with those of transcriptome sequencing, which further verified the reliability of transcriptome sequencing results. (7) With the comprehensive effects of those factors, ancient Camellia oleifera leaves still kept an exuberant vitality, which had a high reference value for the stress resistance biological study and genetic improvement research of Camellia oleifera. References Lim, P. O., Kim, H. J. & Nam, H. G. Leaf Senescence. Annu Rev Plant Biol. 58 , 115–136 (2007). Chen, Y. Z. & Dong, H. Z. Mechanisms and regulation of senescence and maturity performance in cotton. Field Crops Research. 189 , 1–9 (2016). Balazadeh, S. et al. A gene regulatory network controlled by the NAC transcription factor ANAC092/AtNAC2/ORE1 during salt-promoted senescence. Plant J. 62 , 250–264 (2010). Watanabe, M. et al. Comprehensive Dissection of Spatiotemporal Metabolic Shifts in Primary, Secondary, and Lipid Metabolism during Developmental Senescence in Arabidopsis. Plant Physiol. 162 , 1290–1310 (2013). Nooden, L. & Guiamet, J. & John I. Senescence mechanisms. Physiology Planta. 101 , 746–753 (1997). Lee, R. & Chen, S. Programmed cell death during rice leaf senescence is nonapoptotic. New Phytol. 155 , 25–32 (2002). Chang, E. M. et al. De novo Characterization of the Platycladus orientalis Transcriptome and Analysis of Photosynthesis-Related Genes during Aging. Forests. 10 , 393 (2019). Li, Z. et al. In Vitro Propagation of Camellia oleifera Abel. Using Hypocotyl, Cotyledonary Node, and Radicle Explants. HortScience. 51 , 416–421 (2016). Zhang, Y. Y. et al. Circadian Evening Complex represses Jasmonate-induced leaf senescence in, Arabidopsis. Mol Plant. 11 , 326–337 (2018). Liu, P. et al. The Histone H3K4 Demethylase JMJ16 Represses Leaf Senescence in Arabidopsis. Plant Cell. 31 , 430–443 (2019). Zhang, Z. S. et al. Characterization of Photosynthetic Performance during Senescence in Stay-Green and Quick-Leaf-Senescence Zea mays L. Inbred Lines. PLoS One. 7 , e42936 (2012). Chen, D. Q., Wang, S. W., Xiong, B. L., Cao, B. B. & Deng, X. P. Carbon/Nitrogen Imbalance Associated with Drought-Induced Leaf Senescence in Sorghum bicolor. PLoS One. 10 , e0137026 (2015). Bresson, J., Bieker, S., Riester, L., Doll, J. & Zentgraf, U. A guideline for leaf senescence analyses: from quantification to physiological and molecular investigations. J Exp Bot. 69 , 769–786 (2018). Wu, A. H. et al. JUNGBRUNNEN1, a reactive oxygen species-responsive NAC transcription factor, regulates longevity in Arabidopsis. Plant Cell. 24 , 482–506 (2012). Jakhar, S. & Mukherjee, D. Chloroplast pigments, proteins, lipid peroxidation and activities of antioxidative enzymes during maturation and senescence of leaves and reproductive organs of Cajanus cajan L. Physiol Mol Biol Plants. 20 , 171–180 (2014). Park, S. Y. et al. The Senescence-Induced Staygreen Protein Regulates Chlorophyll Degradation. Plant Cell. 19 , 1649–1664 (2007). Kong, X. Q. et al. Gene Expression Profiles Deciphering Leaf Senescence Variation between Early- and Late-Senescence Cotton Lines. PLoS One. 8 , e69847 (2013). Li, H. W. et al. Comparative changes in the antioxidant system in the flag leaf of early and normally senescing near-isogenic lines of wheat (Triticum aestivum L.). Plant Cell Rep. 33 , 1109–1120 (2014). Wang, F. B. et al. Senescence-specific change in ROS scavenging enzyme activities and regulation of various SOD isozymes to ROS levels in psf mutant rice leaves. Plant Physiol Bioch. 109 , 248–261 (2016). Li, Z. et al. Overexpressing the Sedum alfredii Cu/Zn Superoxide Dismutase Increased Resistance to Oxidative Stress in Transgenic Arabidopsis. Front Plant Sci . 8, 1010( 2017 ). Dong, H. Z., Niu, Y. H., Li, W. J. & Zhang, D. M. Effects of cotton rootstock on endogenous cytokinins and abscisic acid in xylem sap and leaves in relation to leaf senescence. J Exp Bot. 59 , 1295–1304 (2008). Riov, J., Dagan, E., Goren, R. & Yang, S. F. Characterization of abscisic Acid-induced ethylene production in citrus leaf and tomato fruit tissues. Plant Physiol. 92 , 48–53 (1990). Fujii, H. & Zhu, J. K. Arabidopsis mutant deficient in 3 abscisic acid-activated protein kinases reveals critical roles in growth, reproduction, and stress. Proc Natl Acad Sci U S A. 106 , 8380–8385 (2009). Umezawa, T. et al. CYP707A3, a major ABA 8 - hydroxylase involved in dehydration andrehydration response in Arabidopsis thaliana. Plant J. 46 , 171–182 (2006). Zhang, Y., Liang, C. Y., Xu, Y., Gianfagna, T. & Huang, B. R. Effects of ipt Gene Expression on Leaf Senescence Induced by Nitrogen or Phosphorus Deficiency in Creeping Bentgrass. J Am Soc Hortic Sci. 35 , 108–115 (2010). Wingler, A., Schaewen, A. V., Leegood, R. C., Lea, P. J. & Quick, W. P. Regulation of leaf senescence by cytokinin, sugars, and light. Effects on NADH-dependent hydroxypyruvate reductase. Plant Physiol. 116 , 329–335 (1998). Xu, Y., Gianfagna, T. & Huang, B. R. Proteomic changes associated with expression of a gene (ipt) controlling cytokinin synthesis for improving heat tolerance in a perennial grass species. J Exp Bot. 61 , 3273–3289 (2010). Le, D. T. et al. Identification and Expression Analysis of Cytokinin Metabolic Genes in Soybean under Normal and Drought Conditions in Relation to Cytokinin Levels. PLoS One. 7 , e42411 (2012). Gao, X. M. & Guo, Y. F. Research Progress of Leaf Senescence Related Genes in Tobacco. J Plant Sci. 6 , 87–92 (2018). Kim, J. I. et al. YUCCA6 over-expression demonstrates auxin function in delaying leaf senescence in Arabidopsis thaliana. J Exp Bot. 62 , 3981–3992 (2011). Zhang, H. S. & Zhou, C. J. Signal transduction in leaf senescence. Plant Mol Biol. 82 , 539–545 (2013). Lin, M. et al. Global analysis of the Gossypium hirsutum L. Transcriptome during leaf senescence by RNA-SEq. BMC Plant Biol. 15 , 43 (2015). Hou, K., Wu, W. & Gan, S. S. SAUR36, a SMALL AUXIN UP RNA Gene, Is Involved in the Promotion of Leaf Senescence in Arabidopsis. Plant Physiol. 161 , 1002–1009 (2013). Ren, H. & Gray, W. M. SAUR Proteins as Effectors of Hormonal and Environmental Signals in Plant Growth. Mol Plant. 8 , 1153–1164 (2015). Olsson, M. Alteration in lipid composition and antioxidative protection during senescence in drought stressed plants and non-drought stressed plants of Pisum sativum. Plant Physiol Bioch. 33 , 547–553 (1995). Yang, S. D., Seo, P. J., Yoon, H. K. & Park, C. M. The Arabidopsis NAC Transcription Factor VNI2 Integrates Abscisic Acid Signals into Leaf Senescence via the COR/RD Genes. Plant Cell. 23 , 2155–2168 (2011). Gao, F. et al. Isolation and characterization of a novel AP2/EREBP-type transcription factor OsAP211 in Oryza sativa. Biol Plantarum. 53 , 643–649 (2009). Vannini, C. et al. Evaluation of transgenic tomato plants ectopically expressing the rice Osmyb4 gene. Plant Sci. 173 , 231–239 (2007). Dai, X. Y. et al. Overexpression of an R1R2R3 MYB Gene, OsMYB3R-2, Increases Tolerance to Freezing, Drought, and Salt Stress in Transgenic Arabidopsis. Plant Physiol. 143 , 1739–1751 (2007). Ding, Z. H. et al. Transgenic expression of MYB15 confers enhanced sensitivity to abscisic acid and improved drought tolerance in Arabidopsis thaliana. J Genet Genomics. 36 , 17–29 (2009). Guo, Y. & Gan, S. AtMYB2 regulates whole plant senescence by inhibiting cytokinin-mediated branching at late stages of development in Arabidopsis. Plant Physiol. 156 , 1612–1619 (2011). Shukla, P. S., Agarwal, P., Gupta, K. & Agarwal, P. K. Molecular characterization of an MYB transcription factor from a succulent halophyte involved in stress tolerance. AoB Plants. 7 , 054 (2015). Pandey, S. P. & Somssich, I. E. The role of WRKY transcription factors in plant immunity. Plant Physiol. 150 , 1648–1655 (2009). Wan, Y. Q. et al. Identification of the WRKY gene family and functional analysis of two genes in Caragana intermedia. BMC Plant Biol. 18 , 31 (2018). Huang, X. S., Wang, W. & Zhang, Q. A basic helix-loop-helix transcription factor, PtrbHLH, of Poncirus trifoliata confers cold tolerance and modulates peroxidase-mediated scavenging of hydrogen peroxide. Plant Physiol. 162 , 1178–1194 (2013). Liu, J. H Saha, G. et al. Genome-wide identification and characterization of MADS-box family genes related to organ development and stress resistance in Brassica rapa. BMC Genomics. 16 , 178 (2015). Yin, W. C. et al. Tomato (Solanum lycopersicum) MADS-box transcription factor SlMBP8 regulates drought, salt tolerance and stress-related genes. Plant Growth Regul. 83 , 55–68 (2017). Mishra, S. K. et al. In the complex family of heat stress transcription factors, HsfA1 has a unique role as master regulator of thermotolerance in tomato. Genes Dev. 16 , 1555–1567 (2002). Reddy, P. S. et al. Role of Heat Shock Proteins in Improving Heat Stress Tolerance in Crop Plants. Heat Shock Proteins and Plants. 10 , 283–307 (2016). Jacob, P., Hirt, H. & Bendahmane, A. The heat-shock protein/chaperone network and multiple stress resistance. Plant Biotechnol J. 15 , 405–414 (2017). Zang, D. D. et al. An Arabidopsis Zinc Finger Protein Increases Abiotic Stress Tolerance by Regulating Sodium and Potassium Homeostasis, Reactive Oxygen Species Scavenging and Osmotic. Potential. Front Plant Sci. 7 , 1272 (2016). Liu, Q. G., Wang, Z. C., Xu, X. M., Zhang, H. Z. & Li, C. H. Genome-Wide Analysis of C2H2 Zinc-Finger Family Transcription Factors and Their Responses to Abiotic Stresses in Poplar (Populus trichocarpa). PLoS One. 10 , e0134753 (2015). Ordiz, M. J., Barbas, C. F. & Beaehy, R. N. Regulation of transgene expression in plants with polydaetyl zinc finger transcription factors. PNAS. 99 , 13290–13295 (2002). Gupta, S. K., Rai, A. K., Kanwar, S. S. & Sharma, T. R. Comparative analysis of zinc finger proteins involved in plant disease resistance. PLoS One. 7 , e42578 (2012). Zhu, M. K. et al. Basic leucine zipper transcription factor SlbZIP1 mediates salt and drought stress tolerance in tomato. BMC Plant Biol. 18 , 83 (2018). Kurep, J., Wang, S. H., Li, Y. & Smalle, J. Proteasome regulation, plant growth and stress tolerance. Plant Signal Behav. 4 , 924–927 (2009). Ho, H. L. Functional Roles of Plant Protein Kinases in Signal Transduction Pathways during Abiotic and Biotic Stress. J Biodivers Biopros. 2 , 147 (2015). Jeworutzki, E. et al. Early signaling through the Arabidopsis pattern recognition receptors FLS2 and EFR involves Ca-associated opening of plasma membrane anion channels. Plant J. 62 , 367–378 (2010). Lee, S. et al. The small GTPase, nucleolar GTP-binding protein 1 (NOG1), has a novel role in plant innate immunity. Sci Rep. 7 , 9260 (2017). Goff, K. E. & Ramonell, K. M.The role and regulation of receptor-like kinases in plant defense. Gene Regul Syst Bio. 1 , 167–175 (2007). Ye, Y. Y. et al. The role of receptor-like protein kinases (RLKs) in abiotic stress response in plants. Plant Cell Rep. 36 , 235–242 (2017). Chinchilla, D. et al. A flagellin-induced complex of the receptor FLS2 and BAK1 initiates plant defence. Nature. 448 , 497–500 (2007). Singh, K. & Foley, R. C. & Oñate-Sánchez, L.Transcription factors in plant defense and stress responses. Curr Opin Plant Biol. 5 , 430–436 (2002). Koyama, T. et al. A regulatory cascade involving class II ETHYLENE RESPONSE FACTOR transcriptional repressors operates in the progression of leaf senescence. Plant Physiol. 162 , 991–1005 (2013). Mase, K. et al. Ethylene-responsive AP2/ERF transcription factor MACD1 participates in phytotoxin-triggered programmed cell death. Mol Plant Microbe Interact. 26 , 868–879 (2013). Takenaka, Y., Nakano, S., Tamoi, M., Sakuda, S. & Fukamizo, T. Chitinase gene expression in response to environmental stresses in Arabidopsis thaliana: chitinase inhibitor allosamidin enhances stress tolerance. Biosci Biotech Bioch. 73 , 1066–1071 (2009). Kumar, M. et al. Chitinases-Potential Candidates for Enhanced Plant Resistance towards Fungal Pathogens. Agriculture. 8 , 88 (2018). Diaz-Mendoza, M. et al. Plant senescence and proteolysis: two processes with one destiny. Genet Mol Biol. 39 , 329–338 (2016). Cruz, C. M. H. et al. Aspartic protease in leaves of common bean (Phaseolus vulgaris L.) and cowpea (Vigna unguiculata L. Walp): enzymatic activity, gene expression and relation to drought susceptibility. FEBS Letters. 492 , 242–246 (2001). Espinoza, C., Medina, C., Somerville, S. & Arce-Johnson, P. Senescence-associated genes induced during compatible viral interactions with grapevine and Arabidopsis. J Exp Bot. 58 , 3197–3212 (2007). Gupta, S., Garg, V., Kant, C. & Bhatia, S. Genome-wide survey and expression analysis of F-box genes in chickpea. BMC Genomics. 16 , 67 (2015). Chen, Y. et al. The F-Box Protein OsFBK12 Targets OsSAMS1 for Degradation and Affects Pleiotropic Phenotypes, Including Leaf Senescence, in Rice1. Plant Physiol. 163 , 1673–1685 (2013). Additional Declarations No competing interests reported. Supplementary Files SupplementaryTable1Differentialgenestatistics.xls SupplementaryTable2Differentialgeneannotation.xlsx Cite Share Download PDF Status: Posted Version 1 posted 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. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-493148","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":25600897,"identity":"c0670eb7-8c6b-4ff6-ae75-ee1bca8ea3a8","order_by":0,"name":"Yongzhong Chen","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Yongzhong","middleName":"","lastName":"Chen","suffix":""},{"id":25600898,"identity":"1f9505a1-a58d-4a5d-bf6b-0152995ce3a9","order_by":1,"name":"Jianjun Chen","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAp0lEQVRIiWNgGAWjYBACxoYDbEDKhoG9HUjxkKAljYHnMLFagACk5TAJWpgbDz978HHH+cQeZgbGB2/biHLYMXPDmWdug7QwG84lTssZNmnettuJ+5kZQAxitfxtOweyhf038VoY2w6AtLAxE6nlmJlkb1uycQ8zY7PknHNEaDGccfiZxM82O9ke9uaDH96UEaXlAMJCItQDgTw/kQpHwSgYBaNgBAMAsqY1mzq5nRUAAAAASUVORK5CYII=","orcid":"","institution":"University of Florida","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Jianjun","middleName":"","lastName":"Chen","suffix":""},{"id":25600899,"identity":"be4a81b4-3b9e-462b-b9b3-cea6fae4c770","order_by":2,"name":"Zhen Zhang","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Zhen","middleName":"","lastName":"Zhang","suffix":""},{"id":25600900,"identity":"dce5f34f-294f-4d78-8f7b-614ffd1b1d41","order_by":3,"name":"Yanming Xu","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Yanming","middleName":"","lastName":"Xu","suffix":""},{"id":25600901,"identity":"e68e6bc7-4936-4df5-9733-40244563f7fb","order_by":4,"name":"Zhilong He","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Zhilong","middleName":"","lastName":"He","suffix":""},{"id":25600902,"identity":"df7741c1-0c66-46dd-a5f0-eec019d79721","order_by":5,"name":"Caixia Liu","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Caixia","middleName":"","lastName":"Liu","suffix":""},{"id":25600903,"identity":"0dea8dd9-a75b-4516-b0a7-a1ca34183034","order_by":6,"name":"Rui Wang","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Rui","middleName":"","lastName":"Wang","suffix":""},{"id":25600904,"identity":"e85fb941-da68-463a-a7b1-e85c8dadc6ea","order_by":7,"name":"Xiangnan Wang","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Xiangnan","middleName":"","lastName":"Wang","suffix":""},{"id":25600905,"identity":"70a6b92f-e412-46fa-b3a4-052dbea77e19","order_by":8,"name":"Yinghe Peng","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Yinghe","middleName":"","lastName":"Peng","suffix":""},{"id":25600906,"identity":"b77824ff-ef5f-4df6-8ef5-cc23b39ef664","order_by":9,"name":"Shaofeng Peng","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Shaofeng","middleName":"","lastName":"Peng","suffix":""},{"id":25600907,"identity":"db6c77b4-9c30-4376-b31a-2e7719803bdd","order_by":10,"name":"Longshen Chen","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Longshen","middleName":"","lastName":"Chen","suffix":""},{"id":25600908,"identity":"5363443e-7fbe-4ca8-b0f0-f2d863994551","order_by":11,"name":"Li Ma","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Ma","suffix":""},{"id":25600909,"identity":"9de5c420-fa54-4884-ab4d-ec362fa7af5c","order_by":12,"name":"Zhigang Li","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Zhigang","middleName":"","lastName":"Li","suffix":""},{"id":25600910,"identity":"4f0ae9b7-56da-4c54-9df5-ca9853b14996","order_by":13,"name":"Wei Tang","email":"","orcid":"","institution":"Hunan Academy of Forestry","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Wei","middleName":"","lastName":"Tang","suffix":""}],"badges":[],"createdAt":"2021-05-04 16:44:08","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-493148/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-493148/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":9067193,"identity":"b5726186-2aeb-42a2-9419-82b87e41eb75","added_by":"auto","created_at":"2021-05-11 21:45:57","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1133375,"visible":true,"origin":"","legend":"The growth status of Camellia oleifera trees with different ages\nNote: Those from left to night were ancient Camellia oleifera (\u003e 100 a), mature tree (30 a).","description":"","filename":"fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-493148/v1/29fdefed74a78a432bbdf769.png"},{"id":9067060,"identity":"e62fd070-0a27-4d15-8feb-fa0d5d74d62c","added_by":"auto","created_at":"2021-05-11 21:39:57","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":22434,"visible":true,"origin":"","legend":"RT-qPCR was used to detect the expression of five differential genes\nNote: P1 is ancient Camellia oleifera (\u003e 100 a), P2 is mature tree (30 a).","description":"","filename":"fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-493148/v1/d9db0df0bfa7263ed37d0dcc.png"},{"id":15673291,"identity":"25431ff9-49cd-4cb4-aa7e-76c709b78a60","added_by":"auto","created_at":"2021-11-18 14:17:14","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1445991,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-493148/v1/f4040dc4-4621-41a8-89bd-75b5a6464f3d.pdf"},{"id":9067127,"identity":"719b2e9c-1739-4760-b029-c1a8780b10cf","added_by":"auto","created_at":"2021-05-11 21:42:57","extension":"xls","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":19456,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable1Differentialgenestatistics.xls","url":"https://assets-eu.researchsquare.com/files/rs-493148/v1/daa73cddbe670a0bacc9bdb8.xls"},{"id":9067125,"identity":"6a6ca246-66e8-45c2-8231-4f70003561b9","added_by":"auto","created_at":"2021-05-11 21:42:57","extension":"xlsx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":42310,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable2Differentialgeneannotation.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-493148/v1/282dde08b179d1408f7f56c1.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eStudy on the Anti-Aging Physiological Characteristics and Molecular Mechanism of \u003cem\u003eCamellia Oleifera\u003c/em\u003e\u003c/p\u003e","fulltext":[{"header":"Introduction","content":" \u003cp\u003eTree senescence refers to the process of decline occurring in plants or some organs during growth and development, which leads to the natural termination of life activities\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e,\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e. Usually, during tree growth and development, the changes of internal factors, such as hormones, metabolic levels, aging-related genes, transcription factors and the effects of external environment factors, such as drought, high temperature, pests may lead to chlorophyll degradation, protein, lipids, nucleic acid, starch and other biological macromolecules hydrolysis\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e,\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e, and cause, accelerate aging, and even cause tree death. At present, the research on the mechanism of tree senescence includes stomatal regulation theory, senescence gene regulation theory, active oxidation theory, nutrition deficit theory, plant hormone regulation theory, external stress and pest theory\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e. The commonness of these theories is to acknowledge that plant senescence and death are the universal laws of life development, and are an active and necessary process in plant growth and development, morphogenesis and environmental response. Moreover, these theories also show that tree senescence is formed under the combined action of many different factors. At the same time, studies have found that different tree senescence processes are different due to different mechanisms and inducing factors of senescence\u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. When encountering many external stress and internal senescence related factors, some plants could form the corresponding resistance as well as the defense mechanism by some antioxidase, the transcription factor, and the signal transduction factor and thus prolonged its senescence process.\u003c/p\u003e \u003cp\u003e \u003cem\u003eCamellia oleifera\u003c/em\u003e which were plants of \u003cem\u003eCamellia\u003c/em\u003e family, \u003cem\u003eCamellia\u003c/em\u003e plants and perennial small trees or shrubs was a specific oleiferous tree species of China which usually distributed at low mountains and hills in South of China\u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eElaeis guineensis\u003c/em\u003e, \u003cem\u003eOlea europaea\u003c/em\u003e, \u003cem\u003eCocos nucleifera\u003c/em\u003e as well as \u003cem\u003eCamellia oleifera\u003c/em\u003e were four edible oleiferous tree species of the world. \u003cem\u003eCamellia oleifera\u003c/em\u003e possessed characteristics of poor soil tolerance, strong resistance, and exuberant vitality. It was common to find the growth of \u003cem\u003eCamellia oleifera\u003c/em\u003e with more than 100 year old age in natural forests. For example, there was a \u003cem\u003eCamellia oleifera\u003c/em\u003e tree with more than 120 year old age grown in Hendong Country, Henyang City in China (N27\u003csup\u003eo\u003c/sup\u003e05\u0026rsquo;, E112\u003csup\u003eo\u003c/sup\u003e56\u0026rsquo;). This single tree produced annual average \u003cem\u003eCamellia\u003c/em\u003e fresh fruit for more than 200 kg during the continuous field observation from 2014 to 2016. What were the factors which maintained the exuberant vitality of those ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e? Nowadays, studies about the transcription level of plants antisenescence mainly focused on the model plants such as \u003cem\u003eArabidopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e,\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e. However, research about antisenescence of \u003cem\u003eCamellia oleifera\u003c/em\u003e was scarce. In this study, focusing on antisenescence of \u003cem\u003eCamellia oleifera\u003c/em\u003e, leaves of \u003cem\u003eCamellia oleifera\u003c/em\u003e with different ages were selected as targets. ROS and antioxidase activities in leaves of mature tree (30 a) and ancient tree (\u0026gt;\u0026thinsp;100 a) were determined. By conducting the transcriptome sequencing analysis, the ROS clearance system of \u003cem\u003eCamellia oleifera\u003c/em\u003e with different ages as well as the expression of differential genes was studied. Furthermore, antisenescence physiological characteristics and molecular mechanism of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e were studied preparing for the further studies of stress resistance molecular biology and genetic improvement.\u003c/p\u003e "},{"header":"Materials And Methods","content":"\u003cdiv\u003e\n\u003ch2\u003eExperimental materials\u003c/h2\u003e\n\u003cp\u003eThe experimental site was located in the experimental forest farm of Hunan Academy of Forestry (east longitude 113\u003csup\u003eo\u003c/sup\u003e01\u003csup\u003e,\u003c/sup\u003e20\u003csup\u003e,,\u003c/sup\u003e, north latitude 28\u003csup\u003eo\u003c/sup\u003e06\u003csup\u003e,\u003c/sup\u003e40\u003csup\u003e,,\u003c/sup\u003e). On April 18th, 2017, good single plant of \u003cem\u003eCamellia oleifera\u003c/em\u003e of mature tree (30 a) and ancient tree (\u0026gt;\u0026thinsp;100 a) with strong growth and good development were sampled (Fig.\u0026nbsp;1). Annual leaves which received consistent illumination, were strong and had no pests and diseases in those single plants at the middle and upper part of the periphery of crown were selected as experimental material, which had three biological replicates. The collected leaves were put into the plastic bags, numbered, quickly put into the ice box and taken back to the laboratory. After rinsing by deionized water and treatment with liquid nitrogen, leaves were put into \u0026minus;\u0026thinsp;70\u003csup\u003eo\u003c/sup\u003eC refrigerator for further analysis.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003cem\u003e\u003cstrong\u003eDetermination methods\u003c/strong\u003e\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eContents of chlorophyll (Chl), malondialdehyde (MDA), hydrogen peroxide (H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e), SOD, POD and CAT were determined by the extraction of acetone, the thiobarbituric acid method, the trichloroacetic acid colorimetry, the riboflavin-NBT photoreduction, the guaiacol method and the permanganate titration method, respectively.\u003c/p\u003e\n\u003cp\u003eSamples were sent to Beijing Genomics Institution in Guangzhou to conduct the transcriptome sequencing. The paired end sequencer of Illumina Hiseq TM 2000 was used for sequencing. Raw data were processed by Beijing Genomics Institution in Guangzhou.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv\u003e\n\u003ch2\u003eData analysis\u003c/h2\u003e\n\u003cp\u003eExcel 2003 was used for the data processing and the figure drawing. SPSS 16.0 was used for data statistics and the \u003cem\u003evariance\u003c/em\u003e analysis. The differential significance of the physiological and biochemical indexes was analyzed by one-way ANOVA test. Multiple comparisons used the least significant difference test (LSD).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv\u003e\n\u003ch2\u003eStatement\u003c/h2\u003e\n\u003cp\u003eWe confirm that the use of plants in the present study complies with international, national and/or institutional guidelines.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAntisenescence \u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003ephysiological indexes analysis of \u003cem\u003eCamellia oleifera\u003c/em\u003e trees with different ages\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThere was a significant difference of chlorophyll content among \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves with different ages. \u003cem\u003eChl \u003c/em\u003ea + Chl b content in leaves of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e was 0.44 mg/g, which was higher than 0.27 mg/g in mature tree. Ratio of Chl a to Chl b in leaves of ancient \u003cem\u003eCamellia oleifera \u003c/em\u003ewere higher than mature tree, which were 10.32、6.34, respectively. MDA and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e contents in leaves of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e were also higher than mature tree, which were 8.83 nmol/g and 126.77 ug/g. Antioxidase activities of SOD, POD and CAT were the highest in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, namely 671.00 (U/g), 22.58 U/(g min) and 498.42 U/(g min), respectively (Table 1). CAT was specific-functional enzyme for the clearance of H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e. POD was also very important for the clearance of surplus H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e in plants. The changing trends of these two enzymes were negatively correlated with the change of H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e to some extent, illustrating that these two could promote the defense of \u003cem\u003eCamellia oleifera\u003c/em\u003e against the attack of H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e and thus prolong senescence. SOD mainly cleared superoxide anion radical (O\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e).\u003c/p\u003e\n\u003cp\u003eIn all, with the increase of the tree age, MDA and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e contents of \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves increased, which resulted in excessive or unbalanced ROS in plants and final senescence. Meantime, \u003cem\u003echlorophyll \u003c/em\u003ein ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves maintained at a relatively higher level. Besides, ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves possessed relatively higher antioxidase activities, which not only effectively guaranteed photosynthesis but also defended the possibly adverse effect caused by the increase of MDA and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e contents. Finally, the normal growth and development was promoted and the senescence process was prolonged.\u003c/p\u003e\n\u003cp\u003eTable 1 Chlorophyll content in \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves of different tree ages\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd width=\"59\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"83\"\u003e\n\u003cp\u003eChl a/Chl b\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"60\"\u003e\n\u003cp\u003eChl a+ Chl b (mg/g)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"70\"\u003e\n\u003cp\u003eMDA (nmol/g)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"61\"\u003e\n\u003cp\u003eH\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e (\u0026mu;g/g)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"61\"\u003e\n\u003cp\u003eSOD\u003c/p\u003e\n\u003cp\u003e(U/g)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"86\"\u003e\n\u003cp\u003ePOD(U/(g\u0026middot;min))\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"88\"\u003e\n\u003cp\u003eCAT(ug/(g\u0026middot;min))\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"59\"\u003e\n\u003cp\u003eAncient tree\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"83\"\u003e\n\u003cp\u003e10.32a(0.44)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"60\"\u003e\n\u003cp\u003e0.44a\u003c/p\u003e\n\u003cp\u003e(0.003)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"70\"\u003e\n\u003cp\u003e8.83a\u003c/p\u003e\n\u003cp\u003e(1.25)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"61\"\u003e\n\u003cp\u003e126.77a\u003c/p\u003e\n\u003cp\u003e(4.75)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"61\"\u003e\n\u003cp\u003e671.00a\u003c/p\u003e\n\u003cp\u003e(35.82)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"86\"\u003e\n\u003cp\u003e22.58a\u003c/p\u003e\n\u003cp\u003e(1.01)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"88\"\u003e\n\u003cp\u003e498.42a\u003c/p\u003e\n\u003cp\u003e(34.45)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"59\"\u003e\n\u003cp\u003eMature tree\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"83\"\u003e\n\u003cp\u003e6.34c(0.20)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"60\"\u003e\n\u003cp\u003e0.27c\u003c/p\u003e\n\u003cp\u003e(0.007)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"70\"\u003e\n\u003cp\u003e6.51ab\u003c/p\u003e\n\u003cp\u003e(0.63)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"61\"\u003e\n\u003cp\u003e109.43b\u003c/p\u003e\n\u003cp\u003e(1.99)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"61\"\u003e\n\u003cp\u003e519.94a\u003c/p\u003e\n\u003cp\u003e(17.56)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"86\"\u003e\n\u003cp\u003e14.34ab\u003c/p\u003e\n\u003cp\u003e(1.00)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"88\"\u003e\n\u003cp\u003e363.80ab\u003c/p\u003e\n\u003cp\u003e(19.40)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eNote: Different letters illustrated the difference of physiological and biochemical indexes among different tree ages (P\u0026lt;0.05). Data in brackets were standard deviation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe differential expression of antisenescence related genes of \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves in stages of different \u003cem\u003etree \u003c/em\u003eages\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe differential expression of chlorophyll degradation related genes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eChlorophyll a/b binding protein gene was very important for the maintenance of the chlorophyll content \u003cem\u003eand \u003c/em\u003ethe photosynthetic ability of the plant. In this study, 10 differentially expressed related genes of chlorophyll a/b-binding protein were found in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, wherein 9 genes were up regulated expressed and 1 gene was down regulated expressed (Table 2). The expression increase of Chlorophyll a/b binding protein gene possibly prolonged the senescence process by enhancing the photosynthetic ability of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e tree.\u003c/p\u003e\n\u003cp\u003eTable 2 Differential expression of chlorophyll a/b-binding protein related genes\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eGene category\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eMain functions\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003eDifferential gene\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003eUp regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eDown regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eChlorophyll a/b-binding protein\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eMaintain chlorophyll content and photosynthetic capacity of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003e9\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe differential expression of antioxidase related genes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSOD, POD, CAT and APX were common antioxidases, which were usually very important for the clearance of surplus ROS in plants and the prolongation of senescence. In this study, 22 antioxidase genes were found significantly differentially expressed in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, wherein 15 antioxidase genes were up regulated expressed and 7 genes were down regulated expressed (Table 3).\u003c/p\u003e\n\u003cp\u003eThese antioxidase genes mainly included SOD, POD, CAT and APX. POD had the most differential genes amount, followed by those of APX, SOD and CAT. The increase of the expression of antioxidase genes could prolong the senescence process by enhancing the antioxidase activity of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e trees.\u003c/p\u003e\n\u003cp\u003eTable 3 Differential expression of antioxidase related genes\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eGene categories\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eMain functions\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"78\"\u003e\n\u003cp\u003eDifferential gene amount\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"135\"\u003e\n\u003cp\u003eUp regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eDown regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eSOD\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eTo clear surplus radicals\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"78\"\u003e\n\u003cp\u003e2\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"135\"\u003e\n\u003cp\u003e0\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e2\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003ePOD\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eTo clear ROS and enhance the stress resistance to drought etc.\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"78\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"135\"\u003e\n\u003cp\u003e9\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eCAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eTo clear surplus H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"78\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"135\"\u003e\n\u003cp\u003e0\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eAPX\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003eTo clear ROS\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"78\"\u003e\n\u003cp\u003e9\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"135\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e3\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eTotal\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"158\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"78\"\u003e\n\u003cp\u003e22\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"135\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e7\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe differential expression of hormone related genes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eHormone levels were closely related to plant senescence. Usually, Auxin and cytokinin (CTK) could prolong plant senescence, whereas abscisic acid (ABA) could accelerate plant senescence. In this study, dramatic differential expression of 40 hormone related genes were found in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, wherein 21 genes were up regulated and 19 genes were down regulated expressed (Table 4). From categories, hormone related genes mainly included ABA, auxin, CKX and SAUR, wherein auxin related differential gene number was the highest, which was followed by that of ABA. Due to the combined effects of those hormone related genes, ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e could maintain hormones at a relatively stable level and further its exuberant vitality.\u003c/p\u003e\n\u003cp\u003eTable 4 Differential expression of hormone related genes\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eGene categories\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"210\"\u003e\n\u003cp\u003eMain functions\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"57\"\u003e\n\u003cp\u003eDifferential gene amount\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"116\"\u003e\n\u003cp\u003eUp regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003eDown regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eABA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"210\"\u003e\n\u003cp\u003eInhibition of plant growth, promotion of leaf abscission and acceleration plant dormancy\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"57\"\u003e\n\u003cp\u003e12\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"116\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eAuxin\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"210\"\u003e\n\u003cp\u003ePromotion of the generation of lateral and adventitious root adjustment of flowering and sex differentiation, adjustment of fruiting and fruit development, and control of apical dominance\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"57\"\u003e\n\u003cp\u003e25\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"116\"\u003e\n\u003cp\u003e13\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003e12\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eCKX\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"210\"\u003e\n\u003cp\u003eMaintenance or reestablishment of the stability and equilibrium of CTK in plants, enhancement of antioxidation ability of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"57\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"116\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003e0\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eSAUR\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"210\"\u003e\n\u003cp\u003eMaintenance of auxin level, adjustment of auxin transport and cell amplification\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"57\"\u003e\n\u003cp\u003e2\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"116\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eTotal\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"210\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"57\"\u003e\n\u003cp\u003e40\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"116\"\u003e\n\u003cp\u003e21\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"100\"\u003e\n\u003cp\u003e19\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe differential expression of stress resistance related genes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe transcription factor, the signal transduction factor, the resistance gene and the defense related gene could significantly enhance the resistance of plants and further effectively answer the external adverse growth factors, respectively. 496 stress resistance related genes were significantly differentially expressed in ancient \u003cem\u003eCamellia oleifera \u003c/em\u003eleaves, wherein 301 genes were up regulated expressed and 195 genes were down regulated expressed. Those stress resistance genes mainly included transcription factors of MYB, NAC, MRKY and zinc Finger etc., signal transduction factors of GTP-binding, receptor protein kinase etc., resistance protein of pathogenesis related protein, disease resistance protein etc., and defense-related gene. The amount of differential genes related to Zinc Finger and disease resistance protein was the highest, followed by other genes of LRR receptor-like serine/threonine-proteinkinase, HSP, MYB, Leucine zipper, bHLH and ERF (Table 5). With the combined effects of those stress resistance related genes, ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e could maintain normal growth through the corresponding adjustment and adaption mechanism to response the external growth stress.\u003c/p\u003e\n\u003cp\u003eTable 5 The differential expression of stress resistance related genes\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eGene categories\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eMain functions\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003eDifferential gene amount\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003eUp regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003eDown regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eZinc Finger\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought, high temperature, salt stress, pathogen\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e128\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e73\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e55\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eHSP\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against high temperature and drought\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e39\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e24\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eMYB\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought, salt, coldness, high temperature stress and enhancement of POD activities of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003ebHLH\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eClearance of ROS and further enhancement of coldness resistance of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e31\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e22\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e9\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eLeucine zipper\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought and salt stress\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e21\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e18\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e3\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eWRKY\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought and salt stress\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e20\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e18\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e2\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eNAC\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought, high temperature, pathogenic bacteria\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e0\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eMADS-box\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought and water stress\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e4\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e3\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e1\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eLRR receptor-like Serine/threonine-proteinkinase\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought and salt stress\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e74\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e41\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e33\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eERF\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eEnhancement of disease resistance of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e21\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e11\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eGTP-binding protein\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eEnhancement of stress resistance and disease resistance of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e19\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e13\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eReceptor protein kinase\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against drought and salt marsh stress\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e8\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e5\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e3\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eDisease resistance protein\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eEnhancement of disease resistance\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e56\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e36\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e20\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003ePathogenesis related protein\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eDefense against various pathogenic bacteria\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e13\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e7\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eChitinase\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eEnhancement of disease resistance of plants\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e8\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e2\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eStress protein\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003eEnhancement of resistance of plants to external stress\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e14\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e7\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e7\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"174\"\u003e\n\u003cp\u003eTotal\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"153\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"84\"\u003e\n\u003cp\u003e496\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"80\"\u003e\n\u003cp\u003e301\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"79\"\u003e\n\u003cp\u003e195\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe differential expression of protein degradation related genes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eProtein degradation which was also an important reason for plant senescence usually occurred due to the function of some proteinase and F-box family protein genes. In this study, 72 functional genes related to protein degradation were found in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves (Table 6). In detail, those included 17 aspartyl protease genes, 11 cysteine proteinase genes and 44 F-box family protein genes. Among cysteine proteinase, 7 genes were up regulated and 4 were down regulated. Among F-box family protein genes and aspartyl protease genes, the expression of most differential genes demonstrated a down regulated trend. With the combined effects, protein degradation of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves may be effectively delayed, which further was possibly helpful for the prolongation of senescence.\u003c/p\u003e\n\u003cp\u003eTable 6 Differential expression of protein degradation related genes\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eGene categories\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eMain functions\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eDifferential gene amount\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003eUp regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eDown regulated gene amount(Ancient tree VS Mature tree)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eF-box\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eInvolvement in protein degradation of plant cells\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e44\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003e17\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e27\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eAspartyl protease\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eInvolvement in senescence and pathogen related protein degradation\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e17\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003e6\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e11\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eCysteine proteinase\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eInvolvement in protein hydrolysis\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e11\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003e7\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e4\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003eTotal\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e72\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"113\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"112\"\u003e\n\u003cp\u003e42\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eVerification of transcriptome differential genes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eReal time quantitative PCR was used to further verify the reliability of transcriptome sequencing results and the expression patterns of differential genes. The selected verification genes include NAC, F-box, aspertyl protection, cystaine protonase, chlorophyll a / B and others. ETIF3H was selected as the internal reference gene. The results showed that the expression of c184012_g3 (F-box)、c178940_g2 (Aspartyl protease)、c163401_g1 (Cysteine proteinase) were down regulated in ancient trees, up regulated in mature tree, c167863_g1 (NAC)、c184170_g1 (Chlorophyll a/b) were up regulated in ancient trees and down regulated in mature tree (Figure 2). In general, the expression patterns of these genes are consistent with the results of transcriptome sequencing, which further verifies the reliability of transcriptome sequencing results.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e"},{"header":"Discussion","content":" \u003cp\u003eThe Chlorophyll content of \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves in ancient trees was significantly higher than those in mature tree. 9 chlorophyll a/b binding proteins in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves demonstrated an up regulated trend. Chlorophyll guaranteed high efficient photosynthesis of plants. Thus, the degradation of chlorophyll was also a main marker of senescence of plant leaves. In \u003cem\u003eZea mays\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e, \u003cem\u003eSorghum bicolor\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e, and \u003cem\u003eArabidopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e, chlorophyll contents were closely related to leaf senescence. Besides, during leaf senescence, some genes involved in photosynthesis would be down regulated expressed, resulting in the decrease of the photosynthetic ability of leaves\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e. Chlorophyll binding protein genes were in a down regulated trend during the senescence process of \u003cem\u003eOryza Sativa\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e and \u003cem\u003eGossypium\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e. In this study, the chlorophyll content and the expression of chlorophyll a/b binding protein gene of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves demonstrated an increasing trend, illustrating that ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves still had a high level of chlorophyll, which was very important for the maintenance of the photosynthesis of the whole tree, the promotion of the growth and development of tree and the prolongation of senescence.\u003c/p\u003e \u003cp\u003eAntioxidase activities of SOD, POD and CAT as well as the expression of POD、APX antioxidase related genes in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves demonstrated an increasing trend. In the senescence process of \u003cem\u003eTriticum aestivum\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e and \u003cem\u003eOryza sativa\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e, activities of SOD, CAT and APX decreased, which will lead to the increase of ROS level and accelerate leaf senescence. A certain concentration of ROS was necessary for the normal physiological process of plants, whereas the excessive ROS would influence and accelerate the senescence process of plants. Thus it was very important to maintain the expression equilibrium of ROS in plants for not only the growth and development but also the prolongation of senescence. Antioxidase and related functional gene expression could effectively clear ROS generated during senescence in leaves\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e,\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e. In this study, multiple antioxidase activities as well as the expression of antioxidase related gene increased in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. Surplus ROS in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves could be cleared in time, which could decrease the cell harm of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e tree from surplus ROS, further maintain normal physiological metabolism of \u003cem\u003eCamellia oleifera\u003c/em\u003e and prolong the senescence of plants.\u003c/p\u003e \u003cp\u003eHormones were very important for the regulation of the growth and development of plants as well as the senescence process of leaves. ABA gene was up regulated expressed in the senescence process of \u003cem\u003eGossypium\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e,\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e, which could influence the senescence of plants by the regulation of growth stress and the induction of ethylene expression\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e,\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e. Abscisic acid 8\u0026rsquo;-hydroxylase which was a key functional enzyme, usually was negatively correlated with ABA content in plants\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e. The increase of CTK level in plants can further prolong and inhibit leaf senescence by ways of decreasing Rubisco content as well as enhancing heat resistance, drought resistance, antioxidation ability and photosynthetic ability of plants\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e,\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e,\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e,\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e,\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e. In this study, the expression of 1 CKX related gene increased in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. In \u003cem\u003eNicotiana tabacum\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e, the increase of the expression of AtCKX could effectively enhance the antioxidation ability of plants. Auxin related genes could prolong plant senescence by inducing auxin contents in plants and adjusting the NAC transcription factors amount\u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e,\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e. In \u003cem\u003eGossypium\u003c/em\u003e, the increase of the expression of auxin related gene could inhibit leaf senescence\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e,\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e. In this study, the expression of 1 SAUR related gene increased in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. SAUR gene played an important role in the maintenance of auxin level, the regulation of auxin transport and cell amplification\u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e,\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e. With the combined effects of those hormone genes, hormone levels in \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves could be maintained in balance, which could promote the benign growth and prolong senescence.\u003c/p\u003e \u003cp\u003eExternal growth stresses of high temperature, low temperature, drought, diseases and pests were usually closely related to plant senescence\u003csup\u003e\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e. Due to many external growth stresses, the transcription factors related to stress in plants would be induced to be expressed. The stress transcription factors could regulate the expression of the stress resistance gene in plants, thus enhancing the stress resistance of plants. In \u003cem\u003eArabisopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e, \u003cem\u003eGossypium\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e, \u003cem\u003eOryza sativa\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e, the increase of the expression of the NAC transcription factor was very useful for prolonging plant senescence, and answering drought as well as salt and marsh stress. The MYB transcription factor expression could enhance the resistance to growth stress of drought, salt, coldness and high temperature in \u003cem\u003eLycopersicon esculentum\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e, \u003cem\u003eArabidopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e,\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e,\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e and \u003cem\u003eSalicornia brachiata\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e. In \u003cem\u003eOryza sativa\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e and \u003cem\u003eArabidopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e, it has been found that WRKY could influence leaf senescence by enhancing the resistance to the adverse factors of growth including drought, high temperature, pathogenic bacteria etc.\u003c/p\u003e \u003cp\u003eIn \u003cem\u003eTrifoliate Orange\u003c/em\u003e, the increase of the expression of the bHLH transcription factor could enhance POD activity of plants, clear ROS, and further enhance coldness resistance of plants\u003csup\u003e\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e\u003c/sup\u003e. In \u003cem\u003eBrassica rapa\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e.and \u003cem\u003eLycopersicon esculentum\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e., it has been found that the increase of the expression of MADS-box gene could enhance the resistance of plants to drought and salt stress. The excessive expression of the HSPs transcription factor was helpful for the enhancement of the stress resistance of plants to high temperature and drought\u003csup\u003e\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e,\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e,\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e\u003c/sup\u003e.. \u003cem\u003eIn Arabidopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e\u003c/sup\u003e. and \u003cem\u003ePopulous Trichocarpa\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e., it has been found that the increase of the expression of the functional gene of Zinc Finger protein could enhance the stress resistance of plants to drought, high temperature, salt stress and pathogen by the adjustment of antioxidase activities of plants\u003csup\u003e\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e,\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e\u003c/sup\u003e.. Zhu \u003cem\u003eet al.\u003c/em\u003e found that the slbzip1 transcription factor of Leucine zipper could play an important role in the resistance of \u003cem\u003eLycopersicon esculentum\u003c/em\u003e to salt and drought by regulating the pathway mediated by ABA\u003csup\u003e\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e\u003c/sup\u003e..\u003c/p\u003e \u003cp\u003eSimilar with the transcription factor, plant received stress stimulation, leaded to the physiological change of plant cells through many signal transduction pathways and finally adapted to the change of external environment\u003csup\u003e\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e,\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e\u003c/sup\u003e.. GTP binding protein was very important for the protection of cell signal transduction and the enhancement of the stress resistance and the disease resistance of plants\u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e,\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e\u003c/sup\u003e. It showed that receptor protein kinase played an important role in growth stress resistance to drought and salt marsh as well as stress responses of antioxidation defense and pathogen reaction\u003csup\u003e\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e,\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e,\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e\u003c/sup\u003e.. LRR receptor-like serine/threonine proteinkinase (FLS2) usually played an important role in the interaction pathway of plant-pathogen, involved in the defense system of pathogen and finally greatly enhanced the disease resistance ability of plants\u003csup\u003e\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e\u003c/sup\u003e.. Ethylene-responsive transcription factors (ERF) not only involved in the plant sescenence and the regulation of many stress response but also took part in the defense and stress answering reaction of plants\u003csup\u003e\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e,\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e,\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e\u003c/sup\u003e..\u003c/p\u003e \u003cp\u003eIn the growth process, plant would not only be influenced by external growth stress but also encountered the invasion of many pathogenic bacteria, which finally exacerbated the senescence process. Chitinase was verified to effectively inhibit activities of some pathogens, intensify the defense system of plants and enhance the disease resistance of plants to many pathogenic bacteria \u003csup\u003e\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e,\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e\u003c/sup\u003e. when encountering stress, plant cells could also increase the defense gene expression of resistance protein, defense-related gene etc. to prevent cells from harm. In ancient trees, the whole gene expression of disease resistance protein and pathogenesis related protein demonstrated an increasing trend.\u003c/p\u003e \u003cp\u003eIn this study, the expression of multiple signal transduction factors, transcription factors and disease resistance related genes were up regulated. With the combined effect of those functional genes, \u003cem\u003eCamellia\u003c/em\u003e oleifera could response to external stress and some pathogen invasion in time, enhance resistance to various growth stresses and pathogenic bacteria through those factors and finally prolong senescence.\u003c/p\u003e \u003cp\u003eThere were many differential genes related to stress resistance of \u003cem\u003eCamellia oleifera\u003c/em\u003e, which was related to the growing environment of \u003cem\u003eCamellia oleifera\u003c/em\u003e itself. \u003cem\u003eCamellia oleifera\u003c/em\u003e was a specific economic tree species in China, which mainly distributed at low mountains and hills in different provinces of South China. In those areas, during the growth process, \u003cem\u003eCamellia oleifera\u003c/em\u003e encountered stresses of low temperature in winter, lots of rain in spring, high temperature in summer and drought. Besides, \u003cem\u003eCamellia oleifera\u003c/em\u003e was susceptible to anthracnose and soft rot. By preliminary speculation, due to the influence of those growth stresses and pathogenic bacteria, the expression of many resistance related genes in \u003cem\u003eCamellia oleifera\u003c/em\u003e including the transcription factors, the resistance factors, the defense factors and the signal transduction genes, which could better defend against those adverse growth factors and finally promote normal growth and development of plants.\u003c/p\u003e \u003cp\u003eIn ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, the overall expression of functional genes related to protein degradation were down regulated. Similar with the situation of chlorophyll degradation, protein would also be degraded during plant senescence\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e. Cysteine proteinase was regarded as the most abundant enzyme which was related to plant leaf senescence\u003csup\u003e\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e\u003c/sup\u003e. and in protein hydrolysis\u003csup\u003e\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e\u003c/sup\u003e. The expression of aspartic protease which took part in petal senescence as well as pathogen related protein degradation\u003csup\u003e\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e\u003c/sup\u003e would be increased during senescence in \u003cem\u003eGlycine max\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e\u003c/sup\u003e and \u003cem\u003eArabidopsis thaliana\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e\u003c/sup\u003e. F-box gene was one of the largest gene families which took part in cell protein degradation in plants. Encountering stresses and during leaf senescence, F-box gene was up regulated expressed in chickpea\u003csup\u003e\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e\u003c/sup\u003e and \u003cem\u003eOryza sativa\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e72\u003c/span\u003e\u003c/sup\u003e. With the degradation of protein, the senescence process of plants would be exacerbated. In ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, the expression of aspartyl protease gene, cysteine proteinase gene and F-box family protein gene were decreased, which effectively inhibited the degradation of protein and thus prolonged senescence.\u003c/p\u003e "},{"header":"Conclusions","content":" \u003cp\u003eThis study shows that: (1) With the increase of the tree age of \u003cem\u003eCamellia oleifera\u003c/em\u003e, the chlorophyll content in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves was significantly higher than those in mature and young trees, which was also verified by the chlorophyll related gene expression in \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. Multiple chlorophyll a/b-binding proteins in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves demonstrated an up regulated trend, illustrating more active vital movement in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves.\u003c/p\u003e \u003cp\u003e(2) The whole expression of related genes of POD, CAT and APX in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves demonstrated an up regulated trend, which was verified by antioxidase contents of SOD, POD and CAT in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, illustrating that the clearance ability of ROS in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves could be enhanced by the increase of antioxidase activities.\u003c/p\u003e \u003cp\u003e(3) The expression of multiple antisenescene hormone related genes increased in \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves, wherein auxin related gene number was the most.\u003c/p\u003e \u003cp\u003e(4) Multiple stress resistance and pathogenic bacteria resistance related genes were found in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. With the combined effects of those genes, \u003cem\u003eCamellia oleifera\u003c/em\u003e could answer various external growth stress and pathogenic bacteria in time and further response correspondingly by the corresponding transcription factor, signal transduction, resistance protein and defense related gene.\u003c/p\u003e \u003cp\u003e(5) The whole expression of the functional differential genes of Aspartyl protease, Cysteine proteinase and F-box family protein, which influenced protein degradation, demonstrated a decreasing trend in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. Thus, protein degradation would be effectively inhibited.\u003c/p\u003e \u003cp\u003e(6) The results of real-time quantitative PCR were consistent with those of transcriptome sequencing, which further verified the reliability of transcriptome sequencing results.\u003c/p\u003e \u003cp\u003e(7) With the comprehensive effects of those factors, ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves still kept an exuberant vitality, which had a high reference value for the stress resistance biological study and genetic improvement research of \u003cem\u003eCamellia oleifera.\u003c/em\u003e\u003c/p\u003e "},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eLim, P. O., Kim, H. J. \u0026amp; Nam, H. G. Leaf Senescence. \u003cem\u003eAnnu Rev Plant Biol.\u003c/em\u003e \u003cb\u003e58\u003c/b\u003e, 115\u0026ndash;136 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen, Y. Z. \u0026amp; Dong, H. Z. Mechanisms and regulation of senescence and maturity performance in cotton. \u003cem\u003eField Crops Research.\u003c/em\u003e \u003cb\u003e189\u003c/b\u003e, 1\u0026ndash;9 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBalazadeh, S. \u003cem\u003eet al.\u003c/em\u003e A gene regulatory network controlled by the NAC transcription factor ANAC092/AtNAC2/ORE1 during salt-promoted senescence. \u003cem\u003ePlant J.\u003c/em\u003e \u003cb\u003e62\u003c/b\u003e, 250\u0026ndash;264 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWatanabe, M. \u003cem\u003eet al.\u003c/em\u003e Comprehensive Dissection of Spatiotemporal Metabolic Shifts in Primary, Secondary, and Lipid Metabolism during Developmental Senescence in Arabidopsis. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e162\u003c/b\u003e, 1290\u0026ndash;1310 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNooden, L. \u0026amp; Guiamet, J. \u0026amp; John I. Senescence mechanisms. \u003cem\u003ePhysiology Planta.\u003c/em\u003e \u003cb\u003e101\u003c/b\u003e, 746\u0026ndash;753 (1997).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee, R. \u0026amp; Chen, S. Programmed cell death during rice leaf senescence is nonapoptotic. \u003cem\u003eNew Phytol.\u003c/em\u003e \u003cb\u003e155\u003c/b\u003e, 25\u0026ndash;32 (2002).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChang, E. M. \u003cem\u003eet al.\u003c/em\u003e De novo Characterization of the Platycladus orientalis Transcriptome and Analysis of Photosynthesis-Related Genes during Aging. \u003cem\u003eForests.\u003c/em\u003e \u003cb\u003e10\u003c/b\u003e, 393 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi, Z. \u003cem\u003eet al.\u003c/em\u003e In Vitro Propagation of Camellia oleifera Abel. Using Hypocotyl, Cotyledonary Node, and Radicle Explants. \u003cem\u003eHortScience.\u003c/em\u003e \u003cb\u003e51\u003c/b\u003e, 416\u0026ndash;421 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang, Y. Y. \u003cem\u003eet al.\u003c/em\u003e Circadian Evening Complex represses Jasmonate-induced leaf senescence in, Arabidopsis. \u003cem\u003eMol Plant.\u003c/em\u003e \u003cb\u003e11\u003c/b\u003e, 326\u0026ndash;337 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLiu, P. \u003cem\u003eet al.\u003c/em\u003e The Histone H3K4 Demethylase JMJ16 Represses Leaf Senescence in Arabidopsis. \u003cem\u003ePlant Cell.\u003c/em\u003e \u003cb\u003e31\u003c/b\u003e, 430\u0026ndash;443 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang, Z. S. \u003cem\u003eet al.\u003c/em\u003e Characterization of Photosynthetic Performance during Senescence in Stay-Green and Quick-Leaf-Senescence Zea mays L. Inbred Lines. \u003cem\u003ePLoS One.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, e42936 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen, D. Q., Wang, S. W., Xiong, B. L., Cao, B. B. \u0026amp; Deng, X. P. Carbon/Nitrogen Imbalance Associated with Drought-Induced Leaf Senescence in Sorghum bicolor. \u003cem\u003ePLoS One.\u003c/em\u003e \u003cb\u003e10\u003c/b\u003e, e0137026 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBresson, J., Bieker, S., Riester, L., Doll, J. \u0026amp; Zentgraf, U. A guideline for leaf senescence analyses: from quantification to physiological and molecular investigations. \u003cem\u003eJ Exp Bot.\u003c/em\u003e \u003cb\u003e69\u003c/b\u003e, 769\u0026ndash;786 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWu, A. H. \u003cem\u003eet al.\u003c/em\u003e JUNGBRUNNEN1, a reactive oxygen species-responsive NAC transcription factor, regulates longevity in Arabidopsis. \u003cem\u003ePlant Cell.\u003c/em\u003e \u003cb\u003e24\u003c/b\u003e, 482\u0026ndash;506 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJakhar, S. \u0026amp; Mukherjee, D. Chloroplast pigments, proteins, lipid peroxidation and activities of antioxidative enzymes during maturation and senescence of leaves and reproductive organs of Cajanus cajan L. \u003cem\u003ePhysiol Mol Biol Plants.\u003c/em\u003e \u003cb\u003e20\u003c/b\u003e, 171\u0026ndash;180 (2014).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePark, S. Y. \u003cem\u003eet al.\u003c/em\u003e The Senescence-Induced Staygreen Protein Regulates Chlorophyll Degradation. \u003cem\u003ePlant Cell.\u003c/em\u003e \u003cb\u003e19\u003c/b\u003e, 1649\u0026ndash;1664 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKong, X. Q. \u003cem\u003eet al.\u003c/em\u003e Gene Expression Profiles Deciphering Leaf Senescence Variation between Early- and Late-Senescence Cotton Lines. \u003cem\u003ePLoS One.\u003c/em\u003e \u003cb\u003e8\u003c/b\u003e, e69847 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi, H. W. \u003cem\u003eet al.\u003c/em\u003e Comparative changes in the antioxidant system in the flag leaf of early and normally senescing near-isogenic lines of wheat (Triticum aestivum L.). \u003cem\u003ePlant Cell Rep.\u003c/em\u003e \u003cb\u003e33\u003c/b\u003e, 1109\u0026ndash;1120 (2014).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang, F. B. \u003cem\u003eet al.\u003c/em\u003e Senescence-specific change in ROS scavenging enzyme activities and regulation of various SOD isozymes to ROS levels in psf mutant rice leaves. \u003cem\u003ePlant Physiol Bioch.\u003c/em\u003e \u003cb\u003e109\u003c/b\u003e, 248\u0026ndash;261 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi, Z. \u003cem\u003eet al.\u003c/em\u003e Overexpressing the Sedum alfredii Cu/Zn Superoxide Dismutase Increased Resistance to Oxidative Stress in Transgenic Arabidopsis.\u003cem\u003eFront Plant Sci\u003c/em\u003e. 8,\u003cem\u003e1010(\u003c/em\u003e 2017\u003cem\u003e).\u003c/em\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDong, H. Z., Niu, Y. H., Li, W. J. \u0026amp; Zhang, D. M. Effects of cotton rootstock on endogenous cytokinins and abscisic acid in xylem sap and leaves in relation to leaf senescence. \u003cem\u003eJ Exp Bot.\u003c/em\u003e \u003cb\u003e59\u003c/b\u003e, 1295\u0026ndash;1304 (2008).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRiov, J., Dagan, E., Goren, R. \u0026amp; Yang, S. F. Characterization of abscisic Acid-induced ethylene production in citrus leaf and tomato fruit tissues. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e92\u003c/b\u003e, 48\u0026ndash;53 (1990).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFujii, H. \u0026amp; Zhu, J. K. Arabidopsis mutant deficient in 3 abscisic acid-activated protein kinases reveals critical roles in growth, reproduction, and stress. \u003cem\u003eProc Natl Acad Sci U S A.\u003c/em\u003e \u003cb\u003e106\u003c/b\u003e, 8380\u0026ndash;8385 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eUmezawa, T. \u003cem\u003eet al.\u003c/em\u003e CYP707A3, a major ABA 8 - hydroxylase involved in dehydration andrehydration response in Arabidopsis thaliana. \u003cem\u003ePlant J.\u003c/em\u003e \u003cb\u003e46\u003c/b\u003e, 171\u0026ndash;182 (2006).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang, Y., Liang, C. Y., Xu, Y., Gianfagna, T. \u0026amp; Huang, B. R. Effects of ipt Gene Expression on Leaf Senescence Induced by Nitrogen or Phosphorus Deficiency in Creeping Bentgrass. \u003cem\u003eJ Am Soc Hortic Sci.\u003c/em\u003e \u003cb\u003e35\u003c/b\u003e, 108\u0026ndash;115 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWingler, A., Schaewen, A. V., Leegood, R. C., Lea, P. J. \u0026amp; Quick, W. P. Regulation of leaf senescence by cytokinin, sugars, and light. Effects on NADH-dependent hydroxypyruvate reductase. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e116\u003c/b\u003e, 329\u0026ndash;335 (1998).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eXu, Y., Gianfagna, T. \u0026amp; Huang, B. R. Proteomic changes associated with expression of a gene (ipt) controlling cytokinin synthesis for improving heat tolerance in a perennial grass species. \u003cem\u003eJ Exp Bot.\u003c/em\u003e \u003cb\u003e61\u003c/b\u003e, 3273\u0026ndash;3289 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLe, D. T. \u003cem\u003eet al.\u003c/em\u003e Identification and Expression Analysis of Cytokinin Metabolic Genes in Soybean under Normal and Drought Conditions in Relation to Cytokinin Levels. \u003cem\u003ePLoS One.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, e42411 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGao, X. M. \u0026amp; Guo, Y. F. Research Progress of Leaf Senescence Related Genes in Tobacco. \u003cem\u003eJ Plant Sci.\u003c/em\u003e \u003cb\u003e6\u003c/b\u003e, 87\u0026ndash;92 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKim, J. I. \u003cem\u003eet al.\u003c/em\u003e YUCCA6 over-expression demonstrates auxin function in delaying leaf senescence in Arabidopsis thaliana. \u003cem\u003eJ Exp Bot.\u003c/em\u003e \u003cb\u003e62\u003c/b\u003e, 3981\u0026ndash;3992 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang, H. S. \u0026amp; Zhou, C. J. Signal transduction in leaf senescence. \u003cem\u003ePlant Mol Biol.\u003c/em\u003e \u003cb\u003e82\u003c/b\u003e, 539\u0026ndash;545 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLin, M. \u003cem\u003eet al.\u003c/em\u003e Global analysis of the Gossypium hirsutum L. Transcriptome during leaf senescence by RNA-SEq. \u003cem\u003eBMC Plant Biol.\u003c/em\u003e \u003cb\u003e15\u003c/b\u003e, 43 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHou, K., Wu, W. \u0026amp; Gan, S. S. SAUR36, a SMALL AUXIN UP RNA Gene, Is Involved in the Promotion of Leaf Senescence in Arabidopsis. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e161\u003c/b\u003e, 1002\u0026ndash;1009 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRen, H. \u0026amp; Gray, W. M. SAUR Proteins as Effectors of Hormonal and Environmental Signals in Plant Growth. \u003cem\u003eMol Plant.\u003c/em\u003e \u003cb\u003e8\u003c/b\u003e, 1153\u0026ndash;1164 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOlsson, M. Alteration in lipid composition and antioxidative protection during senescence in drought stressed plants and non-drought stressed plants of Pisum sativum. \u003cem\u003ePlant Physiol Bioch.\u003c/em\u003e \u003cb\u003e33\u003c/b\u003e, 547\u0026ndash;553 (1995).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYang, S. D., Seo, P. J., Yoon, H. K. \u0026amp; Park, C. M. The Arabidopsis NAC Transcription Factor VNI2 Integrates Abscisic Acid Signals into Leaf Senescence via the COR/RD Genes. \u003cem\u003ePlant Cell.\u003c/em\u003e \u003cb\u003e23\u003c/b\u003e, 2155\u0026ndash;2168 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGao, F. \u003cem\u003eet al.\u003c/em\u003e Isolation and characterization of a novel AP2/EREBP-type transcription factor OsAP211 in Oryza sativa. \u003cem\u003eBiol Plantarum.\u003c/em\u003e \u003cb\u003e53\u003c/b\u003e, 643\u0026ndash;649 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eVannini, C. \u003cem\u003eet al.\u003c/em\u003e Evaluation of transgenic tomato plants ectopically expressing the rice Osmyb4 gene. \u003cem\u003ePlant Sci.\u003c/em\u003e \u003cb\u003e173\u003c/b\u003e, 231\u0026ndash;239 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDai, X. Y. \u003cem\u003eet al.\u003c/em\u003e Overexpression of an R1R2R3 MYB Gene, OsMYB3R-2, Increases Tolerance to Freezing, Drought, and Salt Stress in Transgenic Arabidopsis. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e143\u003c/b\u003e, 1739\u0026ndash;1751 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDing, Z. H. \u003cem\u003eet al.\u003c/em\u003e Transgenic expression of MYB15 confers enhanced sensitivity to abscisic acid and improved drought tolerance in Arabidopsis thaliana. \u003cem\u003eJ Genet Genomics.\u003c/em\u003e \u003cb\u003e36\u003c/b\u003e, 17\u0026ndash;29 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGuo, Y. \u0026amp; Gan, S. AtMYB2 regulates whole plant senescence by inhibiting cytokinin-mediated branching at late stages of development in Arabidopsis. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e156\u003c/b\u003e, 1612\u0026ndash;1619 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShukla, P. S., Agarwal, P., Gupta, K. \u0026amp; Agarwal, P. K. Molecular characterization of an MYB transcription factor from a succulent halophyte involved in stress tolerance. \u003cem\u003eAoB Plants.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, 054 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePandey, S. P. \u0026amp; Somssich, I. E. The role of WRKY transcription factors in plant immunity. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e150\u003c/b\u003e, 1648\u0026ndash;1655 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWan, Y. Q. \u003cem\u003eet al.\u003c/em\u003e Identification of the WRKY gene family and functional analysis of two genes in Caragana intermedia. \u003cem\u003eBMC Plant Biol.\u003c/em\u003e \u003cb\u003e18\u003c/b\u003e, 31 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHuang, X. S., Wang, W. \u0026amp; Zhang, Q. A basic helix-loop-helix transcription factor, PtrbHLH, of Poncirus trifoliata confers cold tolerance and modulates peroxidase-mediated scavenging of hydrogen peroxide. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e162\u003c/b\u003e, 1178\u0026ndash;1194 (2013). Liu, J. H\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSaha, G. \u003cem\u003eet al.\u003c/em\u003e Genome-wide identification and characterization of MADS-box family genes related to organ development and stress resistance in Brassica rapa. \u003cem\u003eBMC Genomics.\u003c/em\u003e \u003cb\u003e16\u003c/b\u003e, 178 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYin, W. C. \u003cem\u003eet al.\u003c/em\u003e Tomato (Solanum lycopersicum) MADS-box transcription factor SlMBP8 regulates drought, salt tolerance and stress-related genes. \u003cem\u003ePlant Growth Regul.\u003c/em\u003e \u003cb\u003e83\u003c/b\u003e, 55\u0026ndash;68 (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMishra, S. K. \u003cem\u003eet al.\u003c/em\u003e In the complex family of heat stress transcription factors, HsfA1 has a unique role as master regulator of thermotolerance in tomato. \u003cem\u003eGenes Dev.\u003c/em\u003e \u003cb\u003e16\u003c/b\u003e, 1555\u0026ndash;1567 (2002).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eReddy, P. S. \u003cem\u003eet al.\u003c/em\u003e Role of Heat Shock Proteins in Improving Heat Stress Tolerance in Crop Plants. \u003cem\u003eHeat Shock Proteins and Plants.\u003c/em\u003e \u003cb\u003e10\u003c/b\u003e, 283\u0026ndash;307 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJacob, P., Hirt, H. \u0026amp; Bendahmane, A. The heat-shock protein/chaperone network and multiple stress resistance. \u003cem\u003ePlant Biotechnol J.\u003c/em\u003e \u003cb\u003e15\u003c/b\u003e, 405\u0026ndash;414 (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZang, D. D. \u003cem\u003eet al.\u003c/em\u003e An Arabidopsis Zinc Finger Protein Increases Abiotic Stress Tolerance by Regulating Sodium and Potassium Homeostasis, Reactive Oxygen Species Scavenging and Osmotic. \u003cem\u003ePotential. Front Plant Sci.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, 1272 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLiu, Q. G., Wang, Z. C., Xu, X. M., Zhang, H. Z. \u0026amp; Li, C. H. Genome-Wide Analysis of C2H2 Zinc-Finger Family Transcription Factors and Their Responses to Abiotic Stresses in Poplar (Populus trichocarpa). \u003cem\u003ePLoS One.\u003c/em\u003e \u003cb\u003e10\u003c/b\u003e, e0134753 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOrdiz, M. J., Barbas, C. F. \u0026amp; Beaehy, R. N. Regulation of transgene expression in plants with polydaetyl zinc finger transcription factors. \u003cem\u003ePNAS.\u003c/em\u003e \u003cb\u003e99\u003c/b\u003e, 13290\u0026ndash;13295 (2002).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGupta, S. K., Rai, A. K., Kanwar, S. S. \u0026amp; Sharma, T. R. Comparative analysis of zinc finger proteins involved in plant disease resistance. \u003cem\u003ePLoS One.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, e42578 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhu, M. K. \u003cem\u003eet al.\u003c/em\u003e Basic leucine zipper transcription factor SlbZIP1 mediates salt and drought stress tolerance in tomato. \u003cem\u003eBMC Plant Biol.\u003c/em\u003e \u003cb\u003e18\u003c/b\u003e, 83 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKurep, J., Wang, S. H., Li, Y. \u0026amp; Smalle, J. Proteasome regulation, plant growth and stress tolerance. \u003cem\u003ePlant Signal Behav.\u003c/em\u003e \u003cb\u003e4\u003c/b\u003e, 924\u0026ndash;927 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHo, H. L. Functional Roles of Plant Protein Kinases in Signal Transduction Pathways during Abiotic and Biotic Stress. \u003cem\u003eJ Biodivers Biopros.\u003c/em\u003e \u003cb\u003e2\u003c/b\u003e, 147 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJeworutzki, E. \u003cem\u003eet al.\u003c/em\u003e Early signaling through the Arabidopsis pattern recognition receptors FLS2 and EFR involves Ca-associated opening of plasma membrane anion channels. \u003cem\u003ePlant J.\u003c/em\u003e \u003cb\u003e62\u003c/b\u003e, 367\u0026ndash;378 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee, S. \u003cem\u003eet al.\u003c/em\u003e The small GTPase, nucleolar GTP-binding protein 1 (NOG1), has a novel role in plant innate immunity. \u003cem\u003eSci Rep.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, 9260 (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGoff, K. E. \u0026amp; Ramonell, K. M.The role and regulation of receptor-like kinases in plant defense. \u003cem\u003eGene Regul Syst Bio.\u003c/em\u003e \u003cb\u003e1\u003c/b\u003e, 167\u0026ndash;175 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYe, Y. Y. \u003cem\u003eet al.\u003c/em\u003e The role of receptor-like protein kinases (RLKs) in abiotic stress response in plants. \u003cem\u003ePlant Cell Rep.\u003c/em\u003e \u003cb\u003e36\u003c/b\u003e, 235\u0026ndash;242 (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChinchilla, D. \u003cem\u003eet al.\u003c/em\u003e A flagellin-induced complex of the receptor FLS2 and BAK1 initiates plant defence. \u003cem\u003eNature.\u003c/em\u003e \u003cb\u003e448\u003c/b\u003e, 497\u0026ndash;500 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSingh, K. \u0026amp; Foley, R. C. \u0026amp; O\u0026ntilde;ate-S\u0026aacute;nchez, L.Transcription factors in plant defense and stress responses. \u003cem\u003eCurr Opin Plant Biol.\u003c/em\u003e \u003cb\u003e5\u003c/b\u003e, 430\u0026ndash;436 (2002).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKoyama, T. \u003cem\u003eet al.\u003c/em\u003e A regulatory cascade involving class II ETHYLENE RESPONSE FACTOR transcriptional repressors operates in the progression of leaf senescence. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e162\u003c/b\u003e, 991\u0026ndash;1005 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMase, K. \u003cem\u003eet al.\u003c/em\u003e Ethylene-responsive AP2/ERF transcription factor MACD1 participates in phytotoxin-triggered programmed cell death. \u003cem\u003eMol Plant Microbe Interact.\u003c/em\u003e \u003cb\u003e26\u003c/b\u003e, 868\u0026ndash;879 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTakenaka, Y., Nakano, S., Tamoi, M., Sakuda, S. \u0026amp; Fukamizo, T. Chitinase gene expression in response to environmental stresses in Arabidopsis thaliana: chitinase inhibitor allosamidin enhances stress tolerance. \u003cem\u003eBiosci Biotech Bioch.\u003c/em\u003e \u003cb\u003e73\u003c/b\u003e, 1066\u0026ndash;1071 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKumar, M. \u003cem\u003eet al.\u003c/em\u003e Chitinases-Potential Candidates for Enhanced Plant Resistance towards Fungal Pathogens. \u003cem\u003eAgriculture.\u003c/em\u003e \u003cb\u003e8\u003c/b\u003e, 88 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDiaz-Mendoza, M. \u003cem\u003eet al.\u003c/em\u003e Plant senescence and proteolysis: two processes with one destiny. \u003cem\u003eGenet Mol Biol.\u003c/em\u003e \u003cb\u003e39\u003c/b\u003e, 329\u0026ndash;338 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCruz, C. M. H. \u003cem\u003eet al.\u003c/em\u003e Aspartic protease in leaves of common bean (Phaseolus vulgaris L.) and cowpea (Vigna unguiculata L. Walp): enzymatic activity, gene expression and relation to drought susceptibility. \u003cem\u003eFEBS Letters.\u003c/em\u003e \u003cb\u003e492\u003c/b\u003e, 242\u0026ndash;246 (2001).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eEspinoza, C., Medina, C., Somerville, S. \u0026amp; Arce-Johnson, P. Senescence-associated genes induced during compatible viral interactions with grapevine and Arabidopsis. \u003cem\u003eJ Exp Bot.\u003c/em\u003e \u003cb\u003e58\u003c/b\u003e, 3197\u0026ndash;3212 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGupta, S., Garg, V., Kant, C. \u0026amp; Bhatia, S. Genome-wide survey and expression analysis of F-box genes in chickpea. \u003cem\u003eBMC Genomics.\u003c/em\u003e \u003cb\u003e16\u003c/b\u003e, 67 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen, Y. \u003cem\u003eet al.\u003c/em\u003e The F-Box Protein OsFBK12 Targets OsSAMS1 for Degradation and Affects Pleiotropic Phenotypes, Including Leaf Senescence, in Rice1. \u003cem\u003ePlant Physiol.\u003c/em\u003e \u003cb\u003e163\u003c/b\u003e, 1673\u0026ndash;1685 (2013).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Camellia oleifera, transduction factor, exuberant vitality, sequencing analysis technique","lastPublishedDoi":"10.21203/rs.3.rs-493148/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-493148/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eTo study the physiological and molecular regulating mechanism of ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e which kept a exuberant vitality for more than one hundred years, leaves of 30a year old and \u0026gt;\u0026thinsp;100 year old \u003cem\u003eCamellia oleifera\u003c/em\u003e were selected as targets. On the basis of the study of the generation and the clearance of reactive oxygen species, sequencing analysis of the transcriptome and expression profiling by high throughput sequencing analysis technique was conducted to study differentially expressed functional genes related to the tree age. It showed that the chlorophyll content and enzyme activities increased in ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves. Expression of chlorophyll a/b binding protein gene, auxin related gene, the signal transduction factor and the transcription factor gene in ancient trees were all higher than mature tree. The down regulated gene expression of inductive genes related to protein degradation in ancient tree. Under the comprehensive function of those factors, ancient \u003cem\u003eCamellia oleifera\u003c/em\u003e leaves still kept an exuberant vitality which was very useful for studies of stress resistance molecular biology and genetic improvement of \u003cem\u003eCamellia oleifera.\u003c/em\u003e\u003c/p\u003e","manuscriptTitle":"Study on the Anti-Aging Physiological Characteristics and Molecular Mechanism of Camellia Oleifera","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2021-05-11 21:39:55","doi":"10.21203/rs.3.rs-493148/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"84fb8caf-7dd0-4b5e-9204-6b23c352b180","owner":[],"postedDate":"May 11th, 2021","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":4245239,"name":"Plant Molecular Biology and Genetics"}],"tags":[],"updatedAt":"2021-08-17T10:49:51+00:00","versionOfRecord":[],"versionCreatedAt":"2021-05-11 21:39:55","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-493148","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-493148","identity":"rs-493148","version":["v1"]},"buildId":"7rjqhiLT3MXkJMwkYKINL","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}