Complete chloroplast genome sequence of Buchanania latifolia (Anacardiaceae): genome structure, and phylogenetic relationships

Complete chloroplast genome sequence of Buchanania latifolia (Anacardiaceae): genome structure, and phylogenetic relationships | 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 Complete chloroplast genome sequence of Buchanania latifolia (Anacardiaceae): genome structure, and phylogenetic relationships Chunmin Mao, Rui Rao, Quwen Lei, WanTing Chen, Liangliang Yue This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4552236/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 The chloroplast (cp) genomes are valuable resource with multiple applications, encompassing species identification, phylogenetic reconstruction, and evolutionary investigations. In this study, the complete chloroplast genome sequence of Buchanania latifolia was de novo sequenced, assembled and annotated. The chloroplast genome of B. latifolia exhibits a typical quadripartite structure, with a total length of 160,088 bp, containing 88 protein-coding sequences (CDS), 37 tRNA genes, and 8 rRNA genes, with an overall GC content of 37.7%. A total of 99 SSR loci and 63 repeat sequences were identified, which can be utilized for marker development, phylogenetic and population studies of B. latifolia . Codon usage analysis revealed a preference for Leu codons ending with A/U. Additionally, the study investigated IR boundaries, DNA polymorphism, positive selection suites, and phylogenetic position. Comparative analysis with five other species from the Anacardiaceae family confirmed the nearly identical and highly conserved chloroplast genome features of B. latifolia , which can be valuable for understanding the plastid evolution and evolutionary relationships within Anacardiaceae. Phylogenetic analysis reveals that B. latifolia is positioned at the base of Anacardiaceae, sister to Choerospondias axillaris , Lannea coromandelica , and Sclerocarya birrea . These findings could provide important genetic information for further research into breeding of Anacardiaceae, phylogeny, and evolution of B. latifolia . Molecular Genetics Plant Molecular Biology and Genetics Buchanania latifolia Roxb. Anacardiaceae Chloroplast genome SSR loci Phylogeny Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Introduction Buchanania latifolia Roxb. also known as Chironji, is a tree species of Anacardiaceae (Weber, 2018 ), native to the eastern Himalayas, Southwest China, and part of Southeast Asia. It's valued for its edible fruits and oil. As a pioneer species, it is commonly found in hot-dry valleys and forests in Yunnan, China. Initially named Coniogeton arborescens , it was reclassified under the genus Buchanania by Blume in 1850 ( Parveen and Ilyas, 2021 ) . The genus Buchanania , with B. latifolia as a notable species, is considered to occupy a basal position in the second ancestral clade of the Anacardiaceae family. This genus comprises approximately 25 species predominantly found in the tropical regions of Asia and Oceania. However, prior to this study, no plastid genome data of the genus Buchanania were available for phylogenetic reconstruction, which could significantly enhance our understanding of phylogenetic relationship, divergence time and biogeographic patterns of these basal clades within the Anacardiaceae family. The next-generation sequencing (NGS) technology is generating vast amounts DNA data including both nuclear and organelle genomes. The chloroplast genome (cp genome), with its self-replication mechanism, relatively independent evolution, appropriate natural evolutionary rate, and unique maternal inheritance is typically highly conserved in terms of genome size, structure, gene content, and gene types in most terrestrial plants (Park and Lee, 2016 , Li et al., 2019a ). These conserved and divergent gene features provide crucial information for identifying, classifying, and reconstructing phylogenetic relationships between species and families (Jansen et al., 2007 ). In this study, the chloroplast genome of B. latifolia was sequenced, assembled, annotated and uploaded to NCBI database. The key features of the chloroplast genome were analyzed, including simple sequence repeat (SSR) loci and optimal codon usage boundaries.A plastid phylogenomic analysis was conducted to examine the phylogenetic relationships among the basal Anacardiaceae taxa. The publication of this plastome will widen our edge of knowledge for the applications both of DNA barcoding and for the biogeographic investigations of this tropical distributed woody clade of Angiosperm. Materials and Methods Plant material, sequencing, and genome assembly Fresh leaves of the Buchanania latifolia were collected from Xinping County, Yunnan Province, China. (102°7′41.75″E, 24°1′54.84′N). The voucher specimen (yue2021006) was deposited in Wetland College of Southwest Forestry University, Kunming, China (email: [email protected] ). Total DNA was extracted from fresh leaves using the E. Z. N. A ® HP Plant DNA Kit (Omega Bio-tek, GA, United States) following the manufacturer’s protocol. Shotgun libraries (350 bp) were constructed using a TruSeq DNA Sample Prep Kit (Illumina, United States). Next-generation sequencing (NGS) was conducted on an Illumina nova-seq 6000 sequencing platform (Illumina, CA, United States), from which approximately 5 Gb of raw NGS data was obtained. Raw reads were filtered using the NGS QC Toolkit (Patel and Jain, 2012) to remove low quality reads and to trim off the sequence adaptors. The filtered reads were then fed into the GetOrganelle (Jin et al., 2020 ) pipeline for de novo genome assembly. The Chloroplast genome of B. latifolia was annotated using Plastid Genome Annotator (PGA) (Qu et al., 2019 ), using the platome of Mangifera indica (GenBank accession number: NC_035239) as a reference. As for the annotated results, the start & stop codons were checked manually, and the intron/exon boundaries of protein-coding genes were checked by using the Geneious Prime (Kearse et al., 2012 ). The chloroplast genome maps were drawn using CPGview ( http://www.1kmpg.cn/cpgview/ ). Finally, the annotated chloroplast genome sequence was submitted to GenBank with an accession number of OM000214. Genome structure analysis The Geneious software was used to export the types and numbers of annotated genes in the cp genome into a CSV file (Kearse et al., 2012 ). Geneious was also used to identify the LSC, SSC, and IR regions in each cp genome sequence and to calculate the AT and GC content. Codon preference analysis The CDS coding sequences of the B. latifolia cp genome were extracted using Geneious (Kearse et al., 2012 ). The number of codons and the relative usage of synonymous codons were analyzed using CodonW 1.4.4 (Peden and John, 2000). Relative Synonymous Codon Usage (RSCU) refers to the relative probability of using a synonymous codon when encoding a specific amino acid. An RSCU > 1 indicates that the codon is used frequently, an RSCU = 1 indicates no preference, and an RSCU < 1 indicates low usage. Repeat sequence analysis Simple sequence repeats (SSRs) were identified and counted using MISA-web (Beier et al., 2017 ), in which the parameters for SSR identification were set at minimal repeat numbers of 10, 5, 4, 3, 3, and 3 for mono-, di-, tri-, tetra-, penta- and hexa- nucleotide repeats, respectively. For large repeats, REPuter (Kurtz et al., 2001 ) was used to identify forward, palindrome, reverse, and complementary long repeats. The hamming distance was set as 3, while the minimal repeat size was 30 bp. Comparative analysis of chloroplast genome IRscope was used to compare IR/SSC and IR/LSC boundaries and junctions among the six Anacardiaceae species ( https://irscope.shinyapps.io/irapp/ ) (Amiryousefi et al., 2018 ). The mVISTA ( https://genome.lbl.gov/vista/mvista/submit.shtml ) (Frazer et al., 2004 ) was used to compare the complete chloroplast genome of the six Anacardiaceae species. Before nucleotide divergence analysis, the cp genome were aligned using MAFFT v7 (Katoh and Standley, 2013 ). The nucleotide variability of the six cp genomes were calculated using DnaSP v5.10 (Librado and Rozas, 2009 ), in which highly mutational hotspot regions were identified through a sliding window analysis. The length of the window frame was set at 4400 bp and a 200 bp step size was selected. Nucleotide substitution rate (Ka/Ks) A total of 79 protein-coding genes shared by the B. latifolia cp genome were extracted using Geneious. After aligning them with MAFFT, the online program ALTER ( https://www.sing-group.org/ALTER/ ) was used to convert the file from ‘.fas’ format to ‘.aln’ format. Subsequently, the ‘.aln’ files were converted to ‘.axt’ format using AXTConvertor from the KaKs-Calculator in a Linux system (Zhang et al., 2006 ). Finally, the ratio of non-synonymous (Ka) to synonymous (Ks) substitutions was calculated using the KaKs-Calculator, selecting the NG model and setting the genetic code type to 11(bacterial and plant plastid codon). When Ka = 0 and Ks = 0, the value of Ka/Ks was represented by NA. Phylogenetic analysis We conducted a phylogenetic analysis of 37 chloroplast genomes in the family Anacardiaceae, using three species from the family Burseraceae, Commiphora gileadensis, Commiphora wightii , and Bursera fagaroides , as outgroups(The accession numbers can be found in Supporting Table 1 ). The Phylogenetic trees of complete chloroplast genomes were constructed using the maximum likelihood (ML) method. ML analysis was conducted using IQ-TREE (Nguyen et al., 2015 ), during which a substitution model of GTR + F + I were automatically calculated and applied with a bootstrap replicates of 1,000 times. Results Genome structure of B. latifolia chloroplast genome The chloroplast (cp) genome of B. latifolia exhibits a quadripartite structure, characterized by a large single-copy (LSC) region, a small single-copy (SSC) region, and a pair of inverted repeats (IRa and IRb), measuring 87,689 bp, 17,941 bp, and 27,217 bp, respectively (refer to Fig. 1 ; Table 1 ). The overall nucleotide composition reveals a predominance of A (30.8%) and T (31.5%), with C (19.2%) and G (18.5%) constituting the remaining nucleotides, resulting in a total AT content of 62.3% and GC content of 37.7%. A comprehensive annotation identified a total of 133 genes within the cp genome of B. latifolia , including 88 protein-coding genes (CDS), 37 transfer RNA genes (tRNAs), and 8 ribosomal RNA genes (rRNAs). Notably, the AT content is higher than the GC content (62.3% vs. 37.7%). Moreover, the GC content varies across different regions, with the LSC, SSC, and IR regions exhibiting values of 35.8%, 32%, and 42.6%, respectively, with the IR regions demonstrating higher GC content compared to the LSC and SSC regions. Functional classification of the genes in the cp genome of B. latifolia categorizes them into four classes: photosynthesis-related genes (44), genes involved in self-replication (74), other annotated genes (6), and genes with unknown functions (7) (see Table 2 ). Table 1 Characteristics of the cp genome of B. latifolia. Category Item Describe Chloroplast genome structure Cp gene/bp 160088 LSC/bp 87713 SSC/bp 17941 IRA/IRB/bp 27217 Gene composition Cp gene 133 CDS 88 tRNA 37 rRNA 8 GC Content (%) Cp gene 37.7 LSC 35.8 SSC 32 IRA/IRB 42.6 A total of 19 duplicated genes including one NADH dehydrogenase subunit gene (ndhB), five self-replication genes ( rpl2 , rpl23 , rps12 , rps19 , rps7 ), four rRNA genes ( rrn16 , rrn23 , rrn4.5 , rrn5 ), seven tRNA genes ( trnA-UGC , trnI-CAU, trnI-GAU , trnL-CAA , trnN-GUU , trnR-ACG , trnV-GAC ), and two unknown function protein genes ( ycf15 , ycf2 ) were illustrated. Most genes are non-coding genes, and a few genes contain one or two introns.Among them, ndhA, ndhB, petB, petD, atpF, rpl16, rpl2, rps12, rps16, rpoC1, trnA-UGC , trnI-GAU , trnK-UUU , trnL-UAA , trnT-CGU , and trnV-UAC contain one intron, while rps12 , clpP and ycf3 contain three introns. It is worth noting that further research using advanced molecular techniques to determine the functions of the 7 genes with unknown functions were necessary (Table 2 ). Table 2 Genes present in chloroplast genome of B. latifolia Category Gene group Gene name Photosynthesis Subunits of photosystem I psaA , psaB , psaC , psaI , psaJ Subunits of photosystem II psbA , psbB , psbC , psbD , psbE , psbF , psbH , psbI , psbJ , psbK , psbL , psbM , psbN , psbT , psbZ Subunits of NADH dehydrogenase ndhA* , ndhB*(2) , ndhC , ndhD , ndhE , ndhF , ndhG , ndhH , ndhI , ndhJ , ndhK Subunits of cytochrome b/f complex petA , petB* , petD* , petG , petL , petN Subunits of ATP synthase atpA , atpB , atpE , atpF* , atpH , atpI Large subunit of rubisco rbcL Subunits photochlorophyllide reductase - Self-replication Proteins of large ribosomal subunit rpl14 , rpl16* , rpl2*(2) , rpl20 , rpl22 , rpl23(2) , rpl32 , rpl33 , rpl36 Proteins of small ribosomal subunit rps11 , rps12**(2) , rps14 , rps15 , rps16* , rps18 , rps19(2) , rps2 , rps3 , rps4, rps7(2) , rps8 Subunits of RNA polymerase rpoA , rpoB , rpoC1* , rpoC2 Ribosomal RNAs rrn16(2), rrn23(2), rrn4.5(2), rrn5(2) Transfer RNAs trnA-UGC*(2) , trnC-GCA , trnD-GUC , trnE-UUC , trnF-GAA , trnG-GCC, trnH-GUG , trnI-CAU(2) , trnI-GAU*(2) , trnK-UUU* , trnL-CAA(2) , trnL-UAA* , trnL-UAG , trnM-CAU , trnN-GUU(2) , trnP-UGG , trnQ-UUG , trnR-ACG(2) , trnR-UCU , trnS-GCU , trnS-GGA , trnS-UGA , trnT-CGU* , trnT-GGU, trnT-UGU , trnV-GAC(2) , trnV-UAC* , trnW-CCA , trnY-GUA , trnfM-CAU Other genes Maturase matK Protease clpP** Envelope membrane protein cemA Acetyl-CoA carboxylase accD c-type cytochrome synthesis gene ccsA Translation initiation factor infA other - Genes of unknown function Conserved hypothetical chloroplast ORF ycf1 , ycf15(2) , ycf2(2) , ycf3** , ycf4 Notes: Gene*: Gene with one introns; Gene**: Gene with two introns; #Gene: Pseudo gene; Gene (2): Number of copies of multi-copy genes. Analysis of codon preference There are significant differences in codon usage patterns among different species. Each amino acid is encoded by at least one codon and up to six codons. This unequal usage of synonymous codons is known as codon bias. Natural selection and base mutations are considered the main factors influencing codon bias. Based on 88 coding sequences (CDS), the codon usage frequency and Relative Synonymous Codon Usage (RSCU) of the cp genome were calculated. These CDS consist of 22,855 codons each, encoding 20 amino acids in the chloroplast genome. Among these, six codons encode arginine (Arg), leucine (Leu), and serine (Ser), while only one codon encodes methionine (Met) and tryptophan (Trp). Among them, leucine (Leu: 10.53%) is the most frequently utilized amino acid, while cysteine (Cys: 1.12%) is the least utilized amino acid in the cp genome. According to RSCU analysis, except for methionine (Met) and tryptophan (Trp), almost all amino acids are encoded by 2–6 synonymous codons. The relative synonymous codon usage for methionine (Met) and tryptophan (Trp) is 1. There were 30 high-frequency codons with RSCU > 1, among which 13 and 16 high-frequency codons ended with A and U, accounting for 93.55% of the total. Only one high-frequency codons ended with G, and no high-frequency codons ending with C were found. A preference of A or U to end codons was indicated for cp genome of the B. latifolia chloroplast. Repeat sequences analysis In this study, 99 SSRs were identified in the cp genome of B. latifolia , including 77 mononucleotides (Mono-), 8 dinucleotides (Di-), 5 trinucleotides (Tri-), 7 tetranucleotides (Tetra-), and 2 hexanucleotides (Hexan-) (Fig. 3 A). 16 SSRs were located in the IR region, 22 SSRs in the SSC region, and the LSC region contained the highest number of SSRs, with 61 (Fig. 3 B). The repeat units were primarily composed of A or T, with B. latifolia cp genome being A/T types rather than G/C types. Furthermore, dinucleotides are primarily composed of AT/AT, with only 1 or 2 occurrences of other nucleotide types.(Fig. 3 C). Within this cp genome, a total of 63 repeat sequences were identified. Among these, forward repeat sequences constituted 39.68% (25 repeats) of the total repeats, while reverse repeat sequences accounted for 3.17% (2 repeats). The remaining 57.14% (36 repeats) were attributed to palindromic repeat sequences. Notably, no complementary repeat sequences were detected (refer to Fig. 4 A). Furthermore, the analysis revealed that repetitive sequences ≤ 40 bp in length were the most prevalent, with 37 occurrences. Subsequently, there were 10 occurrences in the 41–50 bp range, and 16 occurrences in the > 50 bp range (Fig. 4 B). IR expansion and contraction The chloroplast genome of B. latifolia was compared with those of five reported chloroplast genomes from the Anacardiaceae family (Fig. 5 ). The main chloroplast marker genes, rpl22 , ndhF , ycf1 , and trnH , were found at the boundaries of LSC/IRA, IRA/SSC, SSC/IRB, and IRB/LSC, respectively. Among these genes, rpl22 primarily located in the LSC region or crossed the LSC/IRA boundary. In B. latifolia , Dobinea delavayi , Lannea coromandelica , and Pegia nitida , the rpl22 gene was partly located within the IRb region, the IRb part of the rpl22 genes ranged from 39 to 69 bp, while in Searsia paniculata and Semecarpus reticulatus , the entire rpl22 gene was within the IRb region. The ndhF gene was mainly located in the LSC region or across the IRb/SSC boundary, showing positions ranging from 2208 to 2259 within the SSC region. Notably, among all the compared six Anacardiaceae species, the ycf1 gene crossed the IRa/SSC boundary, with positions ranging from 952 to 1479 bp within the IRa region and from 3996 to 4568 bp within the SSC region. The ndhF gene was located within the SSC region, with distances to the IRb/SSC junction of 5, 56, 61, 64, 143, and 154 bp. The trnN gene was entirely located within the IRa region and contracted by 971 to 1479 bp. The trnH gene is located within the LSC region, positioned 74–75 bp away from the IRa/LSC boundary. Structural comparison and divergence hotspot identification analysis Using the annotation of B. latifolia as reference, the chloroplast genome sequences of the five Anacardiaceae species were compared by mVISTA (Fig. 6 ). The sequence divergences remarkably differed among regions. The alignment result indicates that the chloroplast genome is extremely conserved with only few variations detected. The data revealed that the non-coding region was more divergent than coding counterparts. IR regions of all cp genomes were less diverged than the LSC and SSC regions. The rRNA genes were highly conserved comparing to other genes. The exons of the ycf1 gene were regions representing the highest polymorphism. The diversity of nucleic acids can reveal variations in nucleic acid sequences among different species. Regions with high variability can serve as potential molecular markers for population genetics. The nucleotide diversity (Pi) value for the six cp genomes ranged from 0 to 0.14438, with an average of 0.02078. At a cut-off point set at Pi ≥ 0.1, four hypervariable regions, including rps16 (exonl)-trnQ (0.12183), atpF (exon1)-atpH (0.11283), ndhF-rpl32 (0.1098), rpl32-trnL (0.10633), and ycf1 (0.14433), were identified. Among the four hypervariable regions, two were in the LSC region, the other two were in the SSC region. None was detected in the IR region (Fig. 7 ). Adaptive evolution analyses Taking B. latifolia as a reference, alterations in 5 Anacardiaceae cp genomes were examined to uncover patterns of selection among protein coding genes (Table 3 ). Overall, the Ka/Ks ratios of 79 protein-coding genes shared by the cp genome of B. latifolia were compared. The results indicate that only the psbT (1.61) gene in L. coromandelica has a Ka/Ks ratio > 1. Additionally, in S. paniculata , the rpl22 (1.09), rpl32 (1.23), rps16 (1.64) and ycf2 (2.04) genes have Ka/Ks ratios greater than 1, while all other genes have Ka/Ks ratios < 1. Table 3 The 79 protein-coding genes in cp genome of the B. latifolia and 5 Anacardiaceae species were used for ka/ks analysis Gene D. delavayi L. coromandelica P. nitida S. paniculata S. reticulatus Gene D. delavayi L. coromandelica P. nitida S. paniculata S. reticulatus accD 0.40 0.40 0.42 0.33 0.45 psbI 0.00 0.00 0.00 0.00 0.00 atpA 0.07 0.04 0.03 0.03 0.04 psbJ NA NA NA NA NA atpB 0.06 0.07 0.05 0.02 0.10 psbK 0.36 0.68 0.82 0.55 0.18 atpE 0.20 0.00 0.00 0.00 0.00 psbL NA 0.00 0.00 NA 0.00 atpF 0.18 0.21 0.26 0.18 0.35 psbM 0.20 0.20 0.20 0.15 NA atpH 0.00 0.00 0.00 0.00 0.00 psbN 0.00 0.00 0.00 0.00 0.00 atpI 0.04 0.06 0.06 0.06 0.09 psbT 0.52 1.61 0.98 0.73 0.64 ccsA 0.32 0.35 0.27 0.37 0.29 psbZ 0.00 0.17 0.17 0.00 NA cemA 0.22 0.11 0.13 0.65 0.40 rbcL 0.30 0.16 0.17 0.18 0.21 clpP 0.03 0.03 0.04 0.19 0.10 rpl14 0.17 0.25 0.18 0.06 0.08 matK 0.52 0.58 0.48 0.60 0.76 rpl16 0.05 0.05 0.04 0.05 0.10 ndhA 0.23 0.14 0.15 0.23 0.04 rpl2 0.16 0.13 0.11 0.11 0.22 ndhB 0.21 0.16 0.31 0.21 0.16 rpl20 0.63 0.40 0.73 0.22 0.31 ndhC 0.08 0.15 0.22 0.17 0.15 rpl22 0.53 0.50 0.54 1.09 0.83 ndhD 0.16 0.21 0.14 0.17 0.13 rpl23 NA NA NA 0.29 NA ndhE 0.18 0.40 0.30 0.12 0.15 rpl32 0.31 0.56 0.70 1.23 0.78 ndhF 0.22 0.32 0.29 0.26 0.27 rpl33 0.36 0.37 0.28 0.09 0.00 ndhG 0.31 0.24 0.31 0.40 0.19 rpl36 0.13 0.04 0.05 0.20 0.17 ndhH 0.04 0.08 0.06 0.07 0.03 rpoA 0.20 0.14 0.14 0.15 0.25 ndhI 0.24 0.52 0.53 0.27 0.64 rpoB 0.10 0.07 0.11 0.17 0.11 ndhJ 0.11 0.21 0.12 0.56 0.28 rpoC1 0.11 0.18 0.12 0.20 0.05 ndhK 0.11 0.11 0.14 0.35 0.19 rpoC2 0.37 0.35 0.41 0.32 0.31 petA 0.11 0.13 0.17 0.19 0.05 rps11 0.06 0.09 0.10 0.06 0.05 petB 0.03 0.03 0.03 0.03 0.21 rps12 0.18 0.12 0.35 0.00 0.00 petD 0.00 0.00 0.00 0.08 0.00 rps14 0.14 0.20 0.19 0.19 0.07 petG 0.00 NA NA NA 0.00 rps15 0.22 0.47 0.18 0.20 0.86 petL NA NA NA 0.18 0.74 rps16 0.21 0.97 0.65 1.64 0.64 petN NA 0.00 0.32 NA NA rps18 0.10 0.30 0.15 0.15 0.30 psaA 0.06 0.06 0.05 0.08 0.06 rps19 0.00 0.05 0.11 0.00 0.00 psaB 0.02 0.02 0.03 0.01 0.00 rps2 0.09 0.09 0.08 0.16 0.12 psaC 0.00 0.00 0.00 0.00 0.00 rps3 0.25 0.16 0.15 0.14 0.14 psaI 0.15 0.61 0.61 NA NA rps4 0.11 0.17 0.15 0.35 0.24 psaJ 0.33 0.33 NA 0.33 0.33 rps7 0.32 NA NA NA NA psbA 0.02 0.02 0.02 0.02 0.06 rps8 0.16 0.08 0.07 0.07 0.05 psbB 0.03 0.05 0.01 0.08 0.07 ycf1 0.73 0.87 0.99 0.98 0.77 psbC 0.02 0.03 0.01 0.04 0.08 ycf15 0.00 0.00 0.00 0.00 NA psbD 0.00 0.00 0.00 0.00 0.00 ycf2 0.81 0.88 0.83 2.04 0.97 psbE 0.00 0.00 0.00 0.00 0.00 ycf3 0.19 0.29 0.17 0.29 0.29 psbF 0.00 0.00 0.00 0.00 0.00 ycf4 0.21 0.16 0.14 0.42 0.23 psbH 0.04 0.00 0.00 0.00 0.17 Phylogenetic relationships of Anacardiaceae species B. latifolia is positioned within the basal clade of the Anacardiaceae family, sister to the clade containing Choerospondias axillaris , Lannea coromandelica , and Sclerocarya birrea (Fig. 8 ). Buchanania is grouped on the same branch with Choerospondias , Lannea , and Sclerocarya of the Spondioideae. Anacardium and Mangifera are clustered with Semecarpus into the clade of Semecarpeae. Discussion Homology: Feature of Chloroplast Genomes In this study, the chloroplast genome features and phylogenetic relationships of Buchanania latifolia were comprehensively analyzed. The chloroplast genome of B. latifolia showed a high degree of homology compared to other reported chloroplast genomes of Anacardiaceae species, including Dracontomelon delavayi , Lannea coromandelica , Pistacia nitida , Spondias paniculata , and Swintonia reticulata . The genome lengths of these species ranged from 159,485 to 162,509 bp, with a maximum difference of 3,024 bp. The overall GC content is comparable to that of other species in the Anacardiaceae family, such as Rhus chinensis (37.79%), Pistacia weinmannifolia (37.84%), Toxicodendron vernicifluum (37.96%), and Cotinus species (37.9%-38.1%) (Wang et al., 2020 , Zheng et al., 2018 , Liu et al., 2023 ). The number and types of genes were also very similar, reflecting the highly conserved characteristics of chloroplast genomes. The homology among closely related woody species is typically considered reasonable due to their long generation times and the relatively low number of substitutions occurring over a given period. Substitutions from parent plants can only be passed to offspring during the process of germination. The study of codon preference aids in understanding the evolutionary processes of plant species and optimizing the expression of exogenous genes in chloroplasts, enabling the prediction of gene function and expression levels (Li et al., 2019b ). Consistent with previous findings, Buchanania latifolia demonstrates a preference for A or U bases in its chloroplast genome codons, a common trait in plant chloroplast genomes (Zhou et al., 2008 ). In B. latifolia, 30 high-frequency codons were identified, with 29 of them ending with an A or U base, likely influenced by natural selection and mutations (Necşulea and Lobry, 2007 ). Previous studies have also noted that in the chloroplast genomes of Anacardiaceae, high-frequency codons tend to utilize A or U bases as the third codon base (Liu et al., 2023 , Xin et al., 2023 , Wang et al., 2020 ). The most prevalent SSRs in the chloroplast genomes of B. latifolia were mononucleotide repeats. Similar to other plants, chloroplast SSRs predominantly consist of short poly-A or poly-T repeats, with mononucleotide repeats being the most common forms (Tao et al., 2023 , Vu et al., 2020 , Djedid et al., 2021 , Provan et al., 2001 , Yang et al., 2020 ). Moreover, the majority of SSRs are located in the LSC and SSC regions, consistent with previous findings on chloroplast genomes (Alshegaihi, 2024 , Liu et al., 2023 , Wang et al., 2020 ). Palindrome sequences accounted for 39.68%, forward repeated sequences for 57.14%, and reverse sequences for 3.17%. No complementary sequences were found. Forward and palindromic repeats were the most common repeat types, with most dispersed repeats being less than 40 bp, as reported in previous studies (Liang et al., 2020 , Kirov et al., 2020 , Tian et al., 2021 , Yuan et al., 2005 ). The IR region of the chloroplast genome is thought to be the most conservative section. The expansion and contraction of the IR region are pivotal factors influencing the length variation observed in plant chloroplast genomes, typically categorized into two types (Yi et al., 2013 , Zhang et al., 2013 ). Such expansions and contractions in the IR region across most species manifest as minor deviations in the IR/SC boundary within a few fixed genes, which may lead to pseudogenization of certain genes. The examination of the chloroplast genome of B. latifolia and the Anacardiaceae family in this study aligns with prior reports in angiosperms (Liu et al., 2023 , Wang et al., 2020 , Xin et al., 2023 ). Typically, the LSC/IRb boundary is situated on or near rps19 , rpl2 , or rpl22 , while IRb/SSC is generally positioned on ycf1 or between ycf1 and ndhF . The SSC/IRa boundary typically lies on ycf1 , and the IRa/LSC boundary usually falls on or near rps19 , rpl2 , rpl12 , and trnH (Alshegaihi, 2024 ). Intergenic spacers are more divergent than introns and protein-coding sequences (Meng et al., 2018 ). However, pseudogenes suffer the same fate as intergenic spacers due to a lack of functional importance, leading to less conservative strains. Pseudogenization was common in the evolution of chloroplast genomes, such as accD , ccsA , ycf1 , rps19 and psbB pseudogenes (Krawczyk et al., 2018 , Li et al., 2021 ). The most divergent genes among the six Anacardiaceae species were two pseudogenes accD and ycf1 , an intron of the rps16 genes, and the intergenic spacer ndhF-rpl32 . These genes or regions can be utilized for developing molecular markers for species identification and population studies (Magdy et al., 2019 )., The synonymous (Ks) and non-synonymous (Ka) nucleotide substitution pattern is a well-recognized marker for assessing genome evolution; and the Ka/Ks ratio reflects selection pressure on genes (Yang and Nielsen, 2000 , Guo et al., 2017 ). Ka/Ks 1 indicate genes that underwent purifying, neutral, and positive selections, respectively (Yang and Nielsen, 2000 ). Generally, synonymous mutations occur more frequently than nonsynonymous mutations within genes, causing the Ka/Ks values to be below 1 (Makałowski and Boguski, 1998 ). Our results using B. latifolia and five species from the Anacardiaceae family indicate that only the psbT gene in L. coromandelica and the rpl22 , rpl32 , rps16 and ycf2 genes in S. paniculata have Ka/Ks values > 1, suggesting strong positive selection acting on these two genes. Ka/Ks values for all other detected genes are below 1, indicating widespread purifying selection on these chloroplast genomes. Application on Phylogenetic Reconstruction The informative sites provided by single genes or gene combinations are often limited. Differences in the evolutionary trajectories of various gene sequences can result in low resolution of the constructed phylogenetic tree, posing challenges in elucidating the evolutionary relationships within the Anacardiaceae family. To optimize phylogenetic outcomes, we conducted a phylogenomic analysis of Anacardiaceae based on complete chloroplast genomes using the maximum likelihood method. Overall, branch support at the generic level was generally high, with only relatively lower support observed for the branch of the genus Rhus and its sister groups (BS = 68), while support for all other branches was robust (BS = 100). This suggests that our analysis accurately reflects the phylogenetic relationships within Anacardiaceae. B. latifolia is sister to Choerospondias , Lannea , and Sclerocarya , consistent with previous phylogenetic trees constructed using sequences such as nuclear ribosomal external transcribed spacer (ETS), the chloroplast trnL intron and trnL-F intergenic spacer ( trnL-F region), and the chloroplast rps16 intron (Weeks et al., 2014 ). However, Choerospondias , Lannea , and Sclerocarya all belong to the tribe/subfamily Spondiadeae/Spondioideae. Phylogenetically, they are distant from Anacardium and Mangifera , which are in the same tribe/subfamily Anacardieae/Anacardioideae as Buchanania . Although Buchanania is typically considered a member of the tribe/subfamily Anacardieae/Anacardioideae, it is here regarded as a sister group to those traditionally belonging to the tribe/subfamily Spondiadeae/Spondioideae, which aligns with Weeks' view (Weeks et al., 2014 ). Conclusions In this study, the complete chloroplast genome sequence of B. latifolia was assembled and annotated de novo. The chloroplast genome of B. latifolia exhibits a typical quadripartite structure, with a length of 160,088 bp, containing 88 protein-coding sequences (CDS), 37 tRNA genes, and 8 rRNA genes, with a GC content of 37.7%. A total of 99 SSR loci and 63 repeat sequences were identified, which can be utilized for marker development, phylogenetic and population studies of B. latifolia . Codon usage analysis revealed a preference for Leu codons ending with A/U. Additionally, the study investigated IR boundaries, DNA polymorphism, positive selection suites, and phylogenetic position. Comparative analysis with five other species from the Anacardiaceae family confirmed the nearly identical and highly conserved chloroplast genome features of B. latifolia , which can be valuable for understanding the plastid evolution and evolutionary relationships within Anacardiaceae. Phylogenetic analysis reveals that B. latifolia is positioned at the base of Anacardiaceae, sister to Choerospondias , Lannea , and Sclerocarya . These fndings could provide important genetic information for further research into breeding of Anacardiaceae, phylogeny, and evolution of B. latifolia . This study reveals the structural characteristics of the chloroplast genome, SSR loci, and phylogenetic analysis, providing valuable genetic information for understanding the origin and evolution of Anacardiaceae in the future. Declarations Data Availability The genome sequence data that support the fndings of this study are openly available in GenBank of NCBI at https://www.ncbi.nlm.nih.gov/ under the accession OM000214.1. The associated BioProject, SRA, and Bio-Sample numbers are PRJNA783515, SRR17036892, and SAMN23436132 respectively. Funding This research was funded by the National Natural Science Foundation of China (grant No. 42161015). The Key Project of Basic Research of Yunnan Province, China (202202AS07337) to Gao Chen. This work is supported by Yunnan Science and Technology Talent and platform Program (202305AM340008). Acknowledgements Thanks to Zhefei Zeng and Kaiwen Jiang for their support of field sampling. Authors and Affiliations (1) Yunnan Key Laboratory of Plateau Wetland Conservation, Restoration and Ecological Services; National Plateau Wetlands Research Center; Dianchi Lake Ecosystem Observation and Research Station of Yunnan Province; Southwest Forestry University, Kunming, PR, 650224, China Chunmin Mao, Rui Rao, WanTing Chen, Liangliang Yue (2) Technology Centre of Kunming Customs Quwen Lei Contributions LLY designed the study. CMM performed statistical and bioinformatic analyzes and wrote the first draft. RR proofreads the image text and provides suggestions for revision. QWL sampling, data analysis. WTC Molecular Materials Management. CMM and RR contribute equally in this study. All-authors read and approved the final manuscript. Corresponding author Correspondence to Liangliang Yue( [email protected] ) Ethics declarations Ethical approval Not applicable. Informed consent. Not applicable. Conflict of interest The authors declare no competing interests. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4552236","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":312191727,"identity":"1c26ed17-6aef-4f06-930c-08478684e3d4","order_by":0,"name":"Chunmin Mao","email":"","orcid":"","institution":"Southwest Forestry University","correspondingAuthor":false,"prefix":"","firstName":"Chunmin","middleName":"","lastName":"Mao","suffix":""},{"id":312191728,"identity":"71d6fe43-06d1-413d-b24a-c213b0971a10","order_by":1,"name":"Rui Rao","email":"","orcid":"","institution":"Southwest Forestry University","correspondingAuthor":false,"prefix":"","firstName":"Rui","middleName":"","lastName":"Rao","suffix":""},{"id":312191729,"identity":"6d7058b4-28f0-486c-9d30-958fe05b2e9f","order_by":2,"name":"Quwen Lei","email":"","orcid":"","institution":"Southwest Forestry University","correspondingAuthor":false,"prefix":"","firstName":"Quwen","middleName":"","lastName":"Lei","suffix":""},{"id":312191730,"identity":"715263ee-5b3d-4761-abcd-897c7ff8913b","order_by":3,"name":"WanTing Chen","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"WanTing","middleName":"","lastName":"Chen","suffix":""},{"id":312191731,"identity":"b0aad789-00aa-40c5-ab7a-3228fbcf7413","order_by":4,"name":"Liangliang Yue","email":"data:image/png;base64,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","orcid":"","institution":"Southwest Forestry University","correspondingAuthor":true,"prefix":"","firstName":"Liangliang","middleName":"","lastName":"Yue","suffix":""}],"badges":[],"createdAt":"2024-06-09 02:47:15","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":true,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":true},"doi":"10.21203/rs.3.rs-4552236/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4552236/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":58118211,"identity":"4f5e273a-bb14-4b48-a8d0-f8be1784a91e","added_by":"auto","created_at":"2024-06-11 11:24:14","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":777732,"visible":true,"origin":"","legend":"\u003cp\u003eGene map for cp genome of the \u003cem\u003eB. latifolia.\u003c/em\u003e\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/0c127125e559d76b90cf2237.png"},{"id":58117549,"identity":"3be4ecd7-4ada-49fd-bbd0-d1b16e30178d","added_by":"auto","created_at":"2024-06-11 11:16:14","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":79439,"visible":true,"origin":"","legend":"\u003cp\u003eCounting of relative synonymous codon usage (RSCU) of amino acids in the cp genome of \u003cem\u003eB. latifolia\u003c/em\u003e. The colors of the histogram correspond to the colors of the codons.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/455679b11ee481c8740cbda3.png"},{"id":58117550,"identity":"03ae25d0-bea5-47b9-8686-3591a8784eb3","added_by":"auto","created_at":"2024-06-11 11:16:14","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":69883,"visible":true,"origin":"","legend":"\u003cp\u003eAnalysis of chloroplast SSRs. A: Number of SSR types. B: Number of SSRs in different copy regions. C: Number of different SSR repeat unit types.\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/8597660067f6fe85dcd08ff6.png"},{"id":58117548,"identity":"8d8d6895-ed52-4440-9ba2-e6ab475a196c","added_by":"auto","created_at":"2024-06-11 11:16:14","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":31202,"visible":true,"origin":"","legend":"\u003cp\u003eAnalysis of chloroplast genome repeat sequences. A: Frequency of four types of repeat sequences. B: Number of tandem copies. Abbreviations: C, complement; F, forward; P, palindrome; R, reverse.\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/99cdf5460c39913a82671d24.png"},{"id":58119318,"identity":"ab3feee6-310d-42f4-a6a2-cbba2fa0bdc4","added_by":"auto","created_at":"2024-06-11 11:40:14","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":218957,"visible":true,"origin":"","legend":"\u003cp\u003eThe borders of the large single-copy (LSC), small single-copy (SSC), and inverted repeat (IR) regions in cp genomes among 6 Anacardiaceae species or varieties. Genes are denoted in colored boxes. The distribution of base lengths (bp) at boundaries are indicated.\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/88e413e52741ffb4940a2e6b.png"},{"id":58117553,"identity":"7c7ee184-3121-40a3-90dc-ef7e904ceb92","added_by":"auto","created_at":"2024-06-11 11:16:14","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":687838,"visible":true,"origin":"","legend":"\u003cp\u003eSequence alignment of eight Phalaenopsis chloroplast genomes using mVISTA. The vertical scale indicates the percentage of identity, ranging from 50 to 100%. The horizontal axis indicated the coordinates within the chloroplast genome. Genome regions were color coded as exon, intron, and conserved non-coding sequences (CNS) and mRNA.\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/f1d6d362c87e5aa4cd96777d.png"},{"id":58118907,"identity":"f9bbedcd-0a52-492d-a743-814cb14c5b6d","added_by":"auto","created_at":"2024-06-11 11:32:14","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":121614,"visible":true,"origin":"","legend":"\u003cp\u003eNucleotide Variety across the cp genomes of the six Anacardiaceae species. X-axis, position of the midpoint of a window; Y-axis, nucleotide diversity of each window.\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/8b6818d2ffe242e671961929.png"},{"id":58117554,"identity":"9dd03b83-ca6a-4892-911b-5c24999f1182","added_by":"auto","created_at":"2024-06-11 11:16:14","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":253081,"visible":true,"origin":"","legend":"\u003cp\u003ePhylogenetic tree reconstruction of Anacardiaceae species based on maximum-likelihood (ML) analysis using the program IQ-Tree v.6.1 (Nguyen et al., 2015) with 1,000 bootstrap replications in 37 complete chloroplast genome sequences. Using three species from the Burseraceae family as outgroups.\u003c/p\u003e","description":"","filename":"8.png","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/f61b0fc8fce16232545c1ea1.png"},{"id":58119824,"identity":"6d8df241-905e-480a-9154-a3cc85788268","added_by":"auto","created_at":"2024-06-11 11:48:15","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3096674,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4552236/v1/0502e331-913a-4cdb-8ad0-8a5f86e3ce2a.pdf"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003eComplete chloroplast genome sequence of \u003cem\u003eBuchanania latifolia \u003c/em\u003e(Anacardiaceae): genome structure, and phylogenetic relationships\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003e \u003cem\u003eBuchanania latifolia\u003c/em\u003e Roxb. also known as Chironji, is a tree species of Anacardiaceae (Weber, \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2018\u003c/span\u003e), native to the eastern Himalayas, Southwest China, and part of Southeast Asia. It's valued for its edible fruits and oil. As a pioneer species, it is commonly found in hot-dry valleys and forests in Yunnan, China. Initially named \u003cem\u003eConiogeton arborescens\u003c/em\u003e, it was reclassified under the genus \u003cem\u003eBuchanania\u003c/em\u003e by Blume in 1850 \u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003e(\u003c/span\u003eParveen and Ilyas, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2021\u003c/span\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003e)\u003c/span\u003e. The genus \u003cem\u003eBuchanania\u003c/em\u003e, with \u003cem\u003eB. latifolia\u003c/em\u003e as a notable species, is considered to occupy a basal position in the second ancestral clade of the Anacardiaceae family. This genus comprises approximately 25 species predominantly found in the tropical regions of Asia and Oceania. However, prior to this study, no plastid genome data of the genus \u003cem\u003eBuchanania\u003c/em\u003e were available for phylogenetic reconstruction, which could significantly enhance our understanding of phylogenetic relationship, divergence time and biogeographic patterns of these basal clades within the Anacardiaceae family.\u003c/p\u003e \u003cp\u003eThe next-generation sequencing (NGS) technology is generating vast amounts DNA data including both nuclear and organelle genomes. The chloroplast genome (cp genome), with its self-replication mechanism, relatively independent evolution, appropriate natural evolutionary rate, and unique maternal inheritance is typically highly conserved in terms of genome size, structure, gene content, and gene types in most terrestrial plants (Park and Lee, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2016\u003c/span\u003e, Li et al., \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2019a\u003c/span\u003e). These conserved and divergent gene features provide crucial information for identifying, classifying, and reconstructing phylogenetic relationships between species and families (Jansen et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). In this study, the chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e was sequenced, assembled, annotated and uploaded to NCBI database. The key features of the chloroplast genome were analyzed, including simple sequence repeat (SSR) loci and optimal codon usage boundaries.A plastid phylogenomic analysis was conducted to examine the phylogenetic relationships among the basal Anacardiaceae taxa. The publication of this plastome will widen our edge of knowledge for the applications both of DNA barcoding and for the biogeographic investigations of this tropical distributed woody clade of Angiosperm.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003ePlant material, sequencing, and genome assembly\u003c/h2\u003e \u003cp\u003eFresh leaves of the \u003cem\u003eBuchanania latifolia\u003c/em\u003e were collected from Xinping County, Yunnan Province, China. (102\u0026deg;7\u0026prime;41.75\u0026Prime;E, 24\u0026deg;1\u0026prime;54.84\u0026prime;N). The voucher specimen (yue2021006) was deposited in Wetland College of Southwest Forestry University, Kunming, China (email: [email protected]). Total DNA was extracted from fresh leaves using the E. Z. N. A \u0026reg; HP Plant DNA Kit (Omega Bio-tek, GA, United States) following the manufacturer\u0026rsquo;s protocol. Shotgun libraries (350 bp) were constructed using a TruSeq DNA Sample Prep Kit (Illumina, United States). Next-generation sequencing (NGS) was conducted on an Illumina nova-seq 6000 sequencing platform (Illumina, CA, United States), from which approximately 5 Gb of raw NGS data was obtained. Raw reads were filtered using the NGS QC Toolkit (Patel and Jain, 2012) to remove low quality reads and to trim off the sequence adaptors. The filtered reads were then fed into the GetOrganelle (Jin et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2020\u003c/span\u003e) pipeline for \u003cem\u003ede novo\u003c/em\u003e genome assembly. The Chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e was annotated using Plastid Genome Annotator (PGA) (Qu et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2019\u003c/span\u003e), using the platome of \u003cem\u003eMangifera indica\u003c/em\u003e (GenBank accession number: NC_035239) as a reference. As for the annotated results, the start \u0026amp; stop codons were checked manually, and the intron/exon boundaries of protein-coding genes were checked by using the Geneious Prime (Kearse et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). The chloroplast genome maps were drawn using CPGview (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.1kmpg.cn/cpgview/\u003c/span\u003e\u003cspan address=\"http://www.1kmpg.cn/cpgview/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e). Finally, the annotated chloroplast genome sequence was submitted to GenBank with an accession number of OM000214.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eGenome structure analysis\u003c/h2\u003e \u003cp\u003eThe Geneious software was used to export the types and numbers of annotated genes in the cp genome into a CSV file (Kearse et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). Geneious was also used to identify the LSC, SSC, and IR regions in each cp genome sequence and to calculate the AT and GC content.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eCodon preference analysis\u003c/h2\u003e \u003cp\u003eThe CDS coding sequences of the \u003cem\u003eB. latifolia\u003c/em\u003e cp genome were extracted using Geneious (Kearse et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). The number of codons and the relative usage of synonymous codons were analyzed using CodonW 1.4.4 (Peden and John, 2000). Relative Synonymous Codon Usage (RSCU) refers to the relative probability of using a synonymous codon when encoding a specific amino acid. An RSCU\u0026thinsp;\u0026gt;\u0026thinsp;1 indicates that the codon is used frequently, an RSCU\u0026thinsp;=\u0026thinsp;1 indicates no preference, and an RSCU\u0026thinsp;\u0026lt;\u0026thinsp;1 indicates low usage.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eRepeat sequence analysis\u003c/h2\u003e \u003cp\u003eSimple sequence repeats (SSRs) were identified and counted using MISA-web (Beier et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), in which the parameters for SSR identification were set at minimal repeat numbers of 10, 5, 4, 3, 3, and 3 for mono-, di-, tri-, tetra-, penta- and hexa- nucleotide repeats, respectively. For large repeats, REPuter (Kurtz et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2001\u003c/span\u003e) was used to identify forward, palindrome, reverse, and complementary long repeats. The hamming distance was set as 3, while the minimal repeat size was 30 bp.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eComparative analysis of chloroplast genome\u003c/h2\u003e \u003cp\u003eIRscope was used to compare IR/SSC and IR/LSC boundaries and junctions among the six Anacardiaceae species (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://irscope.shinyapps.io/irapp/\u003c/span\u003e\u003cspan address=\"https://irscope.shinyapps.io/irapp/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) (Amiryousefi et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). The mVISTA (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://genome.lbl.gov/vista/mvista/submit.shtml\u003c/span\u003e\u003cspan address=\"https://genome.lbl.gov/vista/mvista/submit.shtml\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) (Frazer et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2004\u003c/span\u003e) was used to compare the complete chloroplast genome of the six Anacardiaceae species. Before nucleotide divergence analysis, the cp genome were aligned using MAFFT v7 (Katoh and Standley, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). The nucleotide variability of the six cp genomes were calculated using DnaSP v5.10 (Librado and Rozas, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2009\u003c/span\u003e), in which highly mutational hotspot regions were identified through a sliding window analysis. The length of the window frame was set at 4400 bp and a 200 bp step size was selected.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eNucleotide substitution rate (Ka/Ks)\u003c/h2\u003e \u003cp\u003eA total of 79 protein-coding genes shared by the \u003cem\u003eB. latifolia\u003c/em\u003e cp genome were extracted using Geneious. After aligning them with MAFFT, the online program ALTER (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.sing-group.org/ALTER/\u003c/span\u003e\u003cspan address=\"https://www.sing-group.org/ALTER/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) was used to convert the file from \u0026lsquo;.fas\u0026rsquo; format to \u0026lsquo;.aln\u0026rsquo; format. Subsequently, the \u0026lsquo;.aln\u0026rsquo; files were converted to \u0026lsquo;.axt\u0026rsquo; format using AXTConvertor from the KaKs-Calculator in a Linux system (Zhang et al., \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). Finally, the ratio of non-synonymous (Ka) to synonymous (Ks) substitutions was calculated using the KaKs-Calculator, selecting the NG model and setting the genetic code type to 11(bacterial and plant plastid codon). When Ka\u0026thinsp;=\u0026thinsp;0 and Ks\u0026thinsp;=\u0026thinsp;0, the value of Ka/Ks was represented by NA.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003ePhylogenetic analysis\u003c/h2\u003e \u003cp\u003eWe conducted a phylogenetic analysis of 37 chloroplast genomes in the family Anacardiaceae, using three species from the family Burseraceae, \u003cem\u003eCommiphora gileadensis, Commiphora wightii\u003c/em\u003e, and \u003cem\u003eBursera fagaroides\u003c/em\u003e, as outgroups(The accession numbers can be found in Supporting Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The Phylogenetic trees of complete chloroplast genomes were constructed using the maximum likelihood (ML) method. ML analysis was conducted using IQ-TREE (Nguyen et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), during which a substitution model of GTR\u0026thinsp;+\u0026thinsp;F\u0026thinsp;+\u0026thinsp;I were automatically calculated and applied with a bootstrap replicates of 1,000 times.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003e \u003cb\u003eGenome structure of\u003c/b\u003e \u003cb\u003eB. latifolia\u003c/b\u003e \u003cb\u003echloroplast genome\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe chloroplast (cp) genome of \u003cem\u003eB. latifolia\u003c/em\u003e exhibits a quadripartite structure, characterized by a large single-copy (LSC) region, a small single-copy (SSC) region, and a pair of inverted repeats (IRa and IRb), measuring 87,689 bp, 17,941 bp, and 27,217 bp, respectively (refer to Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The overall nucleotide composition reveals a predominance of A (30.8%) and T (31.5%), with C (19.2%) and G (18.5%) constituting the remaining nucleotides, resulting in a total AT content of 62.3% and GC content of 37.7%. A comprehensive annotation identified a total of 133 genes within the cp genome of \u003cem\u003eB. latifolia\u003c/em\u003e, including 88 protein-coding genes (CDS), 37 transfer RNA genes (tRNAs), and 8 ribosomal RNA genes (rRNAs). Notably, the AT content is higher than the GC content (62.3% vs. 37.7%). Moreover, the GC content varies across different regions, with the LSC, SSC, and IR regions exhibiting values of 35.8%, 32%, and 42.6%, respectively, with the IR regions demonstrating higher GC content compared to the LSC and SSC regions. Functional classification of the genes in the cp genome of \u003cem\u003eB. latifolia\u003c/em\u003e categorizes them into four classes: photosynthesis-related genes (44), genes involved in self-replication (74), other annotated genes (6), and genes with unknown functions (7) (see Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eCharacteristics of the cp genome of \u003cem\u003eB. latifolia.\u003c/em\u003e\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCategory\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eItem\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eDescribe\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eChloroplast genome structure\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCp gene/bp\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e160088\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLSC/bp\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e87713\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSSC/bp\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e17941\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eIRA/IRB/bp\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e27217\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eGene composition\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCp gene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e133\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCDS\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e88\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003etRNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e37\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003erRNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eGC Content (%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCp gene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e37.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLSC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e35.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSSC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e32\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eIRA/IRB\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e42.6\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eA total of 19 duplicated genes including one NADH dehydrogenase subunit gene (ndhB), five self-replication genes (\u003cem\u003erpl2\u003c/em\u003e, \u003cem\u003erpl23\u003c/em\u003e, \u003cem\u003erps12\u003c/em\u003e, \u003cem\u003erps19\u003c/em\u003e, \u003cem\u003erps7\u003c/em\u003e), four rRNA genes (\u003cem\u003errn16\u003c/em\u003e, \u003cem\u003errn23\u003c/em\u003e, \u003cem\u003errn4.5\u003c/em\u003e, \u003cem\u003errn5\u003c/em\u003e), seven tRNA genes (\u003cem\u003etrnA-UGC\u003c/em\u003e, trnI-CAU, \u003cem\u003etrnI-GAU\u003c/em\u003e, \u003cem\u003etrnL-CAA\u003c/em\u003e, \u003cem\u003etrnN-GUU\u003c/em\u003e, \u003cem\u003etrnR-ACG\u003c/em\u003e, \u003cem\u003etrnV-GAC\u003c/em\u003e), and two unknown function protein genes (\u003cem\u003eycf15\u003c/em\u003e, \u003cem\u003eycf2\u003c/em\u003e) were illustrated. Most genes are non-coding genes, and a few genes contain one or two introns.Among them, ndhA, ndhB, petB, petD, atpF, rpl16, rpl2, rps12, rps16, rpoC1, \u003cem\u003etrnA-UGC\u003c/em\u003e, \u003cem\u003etrnI-GAU\u003c/em\u003e, \u003cem\u003etrnK-UUU\u003c/em\u003e, \u003cem\u003etrnL-UAA\u003c/em\u003e, \u003cem\u003etrnT-CGU\u003c/em\u003e, and \u003cem\u003etrnV-UAC\u003c/em\u003e contain one intron, while \u003cem\u003erps12\u003c/em\u003e, \u003cem\u003eclpP\u003c/em\u003e and \u003cem\u003eycf3\u003c/em\u003e contain three introns. It is worth noting that further research using advanced molecular techniques to determine the functions of the 7 genes with unknown functions were necessary (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eGenes present in chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCategory\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGene group\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eGene name\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"6\" rowspan=\"7\"\u003e \u003cp\u003ePhotosynthesis\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits of photosystem I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003epsaA\u003c/em\u003e, \u003cem\u003epsaB\u003c/em\u003e, \u003cem\u003epsaC\u003c/em\u003e, \u003cem\u003epsaI\u003c/em\u003e, \u003cem\u003epsaJ\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits of photosystem II\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003epsbA\u003c/em\u003e, \u003cem\u003epsbB\u003c/em\u003e, \u003cem\u003epsbC\u003c/em\u003e, \u003cem\u003epsbD\u003c/em\u003e, \u003cem\u003epsbE\u003c/em\u003e, \u003cem\u003epsbF\u003c/em\u003e, \u003cem\u003epsbH\u003c/em\u003e, \u003cem\u003epsbI\u003c/em\u003e, \u003cem\u003epsbJ\u003c/em\u003e, \u003cem\u003epsbK\u003c/em\u003e, \u003cem\u003epsbL\u003c/em\u003e, \u003cem\u003epsbM\u003c/em\u003e, \u003cem\u003epsbN\u003c/em\u003e, \u003cem\u003epsbT\u003c/em\u003e, \u003cem\u003epsbZ\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits of NADH dehydrogenase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003endhA*\u003c/em\u003e, \u003cem\u003endhB*(2)\u003c/em\u003e, \u003cem\u003endhC\u003c/em\u003e, \u003cem\u003endhD\u003c/em\u003e, \u003cem\u003endhE\u003c/em\u003e, \u003cem\u003endhF\u003c/em\u003e, \u003cem\u003endhG\u003c/em\u003e, \u003cem\u003endhH\u003c/em\u003e, \u003cem\u003endhI\u003c/em\u003e, \u003cem\u003endhJ\u003c/em\u003e, \u003cem\u003endhK\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits of cytochrome b/f complex\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003epetA\u003c/em\u003e, \u003cem\u003epetB*\u003c/em\u003e, \u003cem\u003epetD*\u003c/em\u003e, \u003cem\u003epetG\u003c/em\u003e, \u003cem\u003epetL\u003c/em\u003e, \u003cem\u003epetN\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits of ATP synthase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eatpA\u003c/em\u003e, \u003cem\u003eatpB\u003c/em\u003e, \u003cem\u003eatpE\u003c/em\u003e, \u003cem\u003eatpF*\u003c/em\u003e, \u003cem\u003eatpH\u003c/em\u003e, \u003cem\u003eatpI\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLarge subunit of rubisco\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003erbcL\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits photochlorophyllide reductase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003eSelf-replication\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eProteins of large ribosomal subunit\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003erpl14\u003c/em\u003e, \u003cem\u003erpl16*\u003c/em\u003e, \u003cem\u003erpl2*(2)\u003c/em\u003e, \u003cem\u003erpl20\u003c/em\u003e, \u003cem\u003erpl22\u003c/em\u003e, \u003cem\u003erpl23(2)\u003c/em\u003e, \u003cem\u003erpl32\u003c/em\u003e, \u003cem\u003erpl33\u003c/em\u003e, \u003cem\u003erpl36\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eProteins of small ribosomal subunit\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003erps11\u003c/em\u003e, \u003cem\u003erps12**(2)\u003c/em\u003e, \u003cem\u003erps14\u003c/em\u003e, \u003cem\u003erps15\u003c/em\u003e, \u003cem\u003erps16*\u003c/em\u003e, \u003cem\u003erps18\u003c/em\u003e, \u003cem\u003erps19(2)\u003c/em\u003e, \u003cem\u003erps2\u003c/em\u003e, \u003cem\u003erps3\u003c/em\u003e, \u003cem\u003erps4, rps7(2)\u003c/em\u003e, \u003cem\u003erps8\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSubunits of RNA polymerase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003erpoA\u003c/em\u003e, \u003cem\u003erpoB\u003c/em\u003e, \u003cem\u003erpoC1*\u003c/em\u003e, \u003cem\u003erpoC2\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eRibosomal RNAs\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003errn16(2), rrn23(2), rrn4.5(2), rrn5(2)\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTransfer RNAs\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003etrnA-UGC*(2)\u003c/em\u003e, \u003cem\u003etrnC-GCA\u003c/em\u003e, \u003cem\u003etrnD-GUC\u003c/em\u003e, \u003cem\u003etrnE-UUC\u003c/em\u003e, \u003cem\u003etrnF-GAA\u003c/em\u003e, \u003cem\u003etrnG-GCC, trnH-GUG\u003c/em\u003e, \u003cem\u003etrnI-CAU(2)\u003c/em\u003e, \u003cem\u003etrnI-GAU*(2)\u003c/em\u003e, \u003cem\u003etrnK-UUU*\u003c/em\u003e, \u003cem\u003etrnL-CAA(2)\u003c/em\u003e, \u003cem\u003etrnL-UAA*\u003c/em\u003e, \u003cem\u003etrnL-UAG\u003c/em\u003e, \u003cem\u003etrnM-CAU\u003c/em\u003e, \u003cem\u003etrnN-GUU(2)\u003c/em\u003e, \u003cem\u003etrnP-UGG\u003c/em\u003e, \u003cem\u003etrnQ-UUG\u003c/em\u003e, \u003cem\u003etrnR-ACG(2)\u003c/em\u003e, \u003cem\u003etrnR-UCU\u003c/em\u003e, \u003cem\u003etrnS-GCU\u003c/em\u003e, \u003cem\u003etrnS-GGA\u003c/em\u003e, \u003cem\u003etrnS-UGA\u003c/em\u003e, \u003cem\u003etrnT-CGU*\u003c/em\u003e, \u003cem\u003etrnT-GGU, trnT-UGU\u003c/em\u003e, \u003cem\u003etrnV-GAC(2)\u003c/em\u003e, \u003cem\u003etrnV-UAC*\u003c/em\u003e, \u003cem\u003etrnW-CCA\u003c/em\u003e, \u003cem\u003etrnY-GUA\u003c/em\u003e, \u003cem\u003etrnfM-CAU\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"6\" rowspan=\"7\"\u003e \u003cp\u003eOther genes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMaturase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003ematK\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eProtease\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eclpP**\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEnvelope membrane protein\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003ecemA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAcetyl-CoA carboxylase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eaccD\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ec-type cytochrome synthesis gene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eccsA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTranslation initiation factor\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003einfA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eother\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003e-\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGenes of unknown function\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eConserved hypothetical chloroplast ORF\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eycf1\u003c/em\u003e, \u003cem\u003eycf15(2)\u003c/em\u003e, \u003cem\u003eycf2(2)\u003c/em\u003e, \u003cem\u003eycf3**\u003c/em\u003e, \u003cem\u003eycf4\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"3\"\u003eNotes: Gene*: Gene with one introns; Gene**: Gene with two introns; #Gene: Pseudo gene; Gene (2): Number of copies of multi-copy genes.\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eAnalysis of codon preference\u003c/h2\u003e \u003cp\u003eThere are significant differences in codon usage patterns among different species. Each amino acid is encoded by at least one codon and up to six codons. This unequal usage of synonymous codons is known as codon bias. Natural selection and base mutations are considered the main factors influencing codon bias. Based on 88 coding sequences (CDS), the codon usage frequency and Relative Synonymous Codon Usage (RSCU) of the cp genome were calculated. These CDS consist of 22,855 codons each, encoding 20 amino acids in the chloroplast genome. Among these, six codons encode arginine (Arg), leucine (Leu), and serine (Ser), while only one codon encodes methionine (Met) and tryptophan (Trp). Among them, leucine (Leu: 10.53%) is the most frequently utilized amino acid, while cysteine (Cys: 1.12%) is the least utilized amino acid in the cp genome. According to RSCU analysis, except for methionine (Met) and tryptophan (Trp), almost all amino acids are encoded by 2\u0026ndash;6 synonymous codons. The relative synonymous codon usage for methionine (Met) and tryptophan (Trp) is 1. There were 30 high-frequency codons with RSCU\u0026thinsp;\u0026gt;\u0026thinsp;1, among which 13 and 16 high-frequency codons ended with A and U, accounting for 93.55% of the total. Only one high-frequency codons ended with G, and no high-frequency codons ending with C were found. A preference of A or U to end codons was indicated for cp genome of the \u003cem\u003eB. latifolia\u003c/em\u003e chloroplast.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eRepeat sequences analysis\u003c/h2\u003e \u003cp\u003eIn this study, 99 SSRs were identified in the cp genome of \u003cem\u003eB. latifolia\u003c/em\u003e, including 77 mononucleotides (Mono-), 8 dinucleotides (Di-), 5 trinucleotides (Tri-), 7 tetranucleotides (Tetra-), and 2 hexanucleotides (Hexan-) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA). 16 SSRs were located in the IR region, 22 SSRs in the SSC region, and the LSC region contained the highest number of SSRs, with 61 (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB). The repeat units were primarily composed of A or T, with \u003cem\u003eB. latifolia\u003c/em\u003e cp genome being A/T types rather than G/C types. Furthermore, dinucleotides are primarily composed of AT/AT, with only 1 or 2 occurrences of other nucleotide types.(Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWithin this cp genome, a total of 63 repeat sequences were identified. Among these, forward repeat sequences constituted 39.68% (25 repeats) of the total repeats, while reverse repeat sequences accounted for 3.17% (2 repeats). The remaining 57.14% (36 repeats) were attributed to palindromic repeat sequences. Notably, no complementary repeat sequences were detected (refer to Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA). Furthermore, the analysis revealed that repetitive sequences\u0026thinsp;\u0026le;\u0026thinsp;40 bp in length were the most prevalent, with 37 occurrences. Subsequently, there were 10 occurrences in the 41\u0026ndash;50 bp range, and 16 occurrences in the \u0026gt;\u0026thinsp;50 bp range (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eB).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eIR expansion and contraction\u003c/h2\u003e \u003cp\u003eThe chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e was compared with those of five reported chloroplast genomes from the Anacardiaceae family (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). The main chloroplast marker genes, \u003cem\u003erpl22\u003c/em\u003e, \u003cem\u003endhF\u003c/em\u003e, \u003cem\u003eycf1\u003c/em\u003e, \u003cem\u003eand trnH\u003c/em\u003e, were found at the boundaries of LSC/IRA, IRA/SSC, SSC/IRB, and IRB/LSC, respectively. Among these genes, rpl22 primarily located in the LSC region or crossed the LSC/IRA boundary. In \u003cem\u003eB. latifolia\u003c/em\u003e, \u003cem\u003eDobinea delavayi\u003c/em\u003e, \u003cem\u003eLannea coromandelica\u003c/em\u003e, and \u003cem\u003ePegia nitida\u003c/em\u003e, the \u003cem\u003erpl22\u003c/em\u003e gene was partly located within the IRb region, the IRb part of the \u003cem\u003erpl22\u003c/em\u003e genes ranged from 39 to 69 bp, while in \u003cem\u003eSearsia paniculata\u003c/em\u003e and \u003cem\u003eSemecarpus reticulatus\u003c/em\u003e, the entire \u003cem\u003erpl22\u003c/em\u003e gene was within the IRb region. The \u003cem\u003endhF\u003c/em\u003e gene was mainly located in the LSC region or across the IRb/SSC boundary, showing positions ranging from 2208 to 2259 within the SSC region. Notably, among all the compared six Anacardiaceae species, the ycf1 gene crossed the IRa/SSC boundary, with positions ranging from 952 to 1479 bp within the IRa region and from 3996 to 4568 bp within the SSC region. The \u003cem\u003endhF\u003c/em\u003e gene was located within the SSC region, with distances to the IRb/SSC junction of 5, 56, 61, 64, 143, and 154 bp. The \u003cem\u003etrnN\u003c/em\u003e gene was entirely located within the IRa region and contracted by 971 to 1479 bp. The \u003cem\u003etrnH\u003c/em\u003e gene is located within the LSC region, positioned 74\u0026ndash;75 bp away from the IRa/LSC boundary.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003eStructural comparison and divergence hotspot identification analysis\u003c/h2\u003e \u003cp\u003eUsing the annotation of \u003cem\u003eB. latifolia\u003c/em\u003e as reference, the chloroplast genome sequences of the five Anacardiaceae species were compared by mVISTA (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e). The sequence divergences remarkably differed among regions. The alignment result indicates that the chloroplast genome is extremely conserved with only few variations detected. The data revealed that the non-coding region was more divergent than coding counterparts. IR regions of all cp genomes were less diverged than the LSC and SSC regions. The rRNA genes were highly conserved comparing to other genes. The exons of the ycf1 gene were regions representing the highest polymorphism.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe diversity of nucleic acids can reveal variations in nucleic acid sequences among different species. Regions with high variability can serve as potential molecular markers for population genetics. The nucleotide diversity (Pi) value for the six cp genomes ranged from 0 to 0.14438, with an average of 0.02078. At a cut-off point set at Pi\u0026thinsp;\u0026ge;\u0026thinsp;0.1, four hypervariable regions, including \u003cem\u003erps16 (exonl)-trnQ\u003c/em\u003e (0.12183), \u003cem\u003eatpF (exon1)-atpH\u003c/em\u003e (0.11283), \u003cem\u003endhF-rpl32\u003c/em\u003e(0.1098), \u003cem\u003erpl32-trnL\u003c/em\u003e (0.10633), and \u003cem\u003eycf1\u003c/em\u003e (0.14433), were identified. Among the four hypervariable regions, two were in the LSC region, the other two were in the SSC region. None was detected in the IR region (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eAdaptive evolution analyses\u003c/h2\u003e \u003cp\u003eTaking \u003cem\u003eB. latifolia\u003c/em\u003e as a reference, alterations in 5 Anacardiaceae cp genomes were examined to uncover patterns of selection among protein coding genes (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Overall, the Ka/Ks ratios of 79 protein-coding genes shared by the cp genome of \u003cem\u003eB. latifolia\u003c/em\u003e were compared. The results indicate that only the \u003cem\u003epsbT\u003c/em\u003e (1.61) gene in \u003cem\u003eL. coromandelica\u003c/em\u003e has a Ka/Ks ratio\u0026thinsp;\u0026gt;\u0026thinsp;1. Additionally, in \u003cem\u003eS. paniculata\u003c/em\u003e, the rpl22 (1.09), \u003cem\u003erpl32\u003c/em\u003e (1.23), \u003cem\u003erps16\u003c/em\u003e (1.64) and \u003cem\u003eycf2\u003c/em\u003e (2.04) genes have Ka/Ks ratios greater than 1, while all other genes have Ka/Ks ratios\u0026thinsp;\u0026lt;\u0026thinsp;1.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe 79 protein-coding genes in cp genome of the B. latifolia and 5 Anacardiaceae species were used for ka/ks analysis\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"12\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGene\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eD. delavayi\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eL. coromandelica\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cem\u003eP. nitida\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cem\u003eS. paniculata\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cem\u003eS. reticulatus\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cem\u003eGene\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cem\u003eD. delavayi\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cem\u003eL. coromandelica\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u003cem\u003eP. nitida\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\"\u003e \u003cp\u003e\u003cem\u003eS. paniculata\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c12\"\u003e \u003cp\u003e\u003cem\u003eS. reticulatus\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eaccD\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.40\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.40\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.42\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.33\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.45\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbI\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eatpA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.07\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbJ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eatpB\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.07\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbK\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.36\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.68\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.82\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.55\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e0.18\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eatpE\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbL\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eatpF\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.26\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eatpH\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbN\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eatpI\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.09\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003epsbT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.52\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.61\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.98\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.73\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e0.64\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eccsA\u003c/em\u003e\u003c/p\u003e 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\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eycf4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.42\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e0.23\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003epsbH\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003ePhylogenetic relationships of Anacardiaceae species\u003c/h2\u003e \u003cp\u003e \u003cem\u003eB. latifolia\u003c/em\u003e is positioned within the basal clade of the Anacardiaceae family, sister to the clade containing \u003cem\u003eChoerospondias axillaris\u003c/em\u003e, \u003cem\u003eLannea coromandelica\u003c/em\u003e, and \u003cem\u003eSclerocarya birrea\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e). \u003cem\u003eBuchanania\u003c/em\u003e is grouped on the same branch with \u003cem\u003eChoerospondias\u003c/em\u003e, \u003cem\u003eLannea\u003c/em\u003e, and \u003cem\u003eSclerocarya\u003c/em\u003e of the Spondioideae. \u003cem\u003eAnacardium\u003c/em\u003e and \u003cem\u003eMangifera\u003c/em\u003e are clustered with \u003cem\u003eSemecarpus\u003c/em\u003e into the clade of Semecarpeae.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003eHomology: Feature of Chloroplast Genomes\u003c/h2\u003e \u003cp\u003eIn this study, the chloroplast genome features and phylogenetic relationships of \u003cem\u003eBuchanania latifolia\u003c/em\u003e were comprehensively analyzed. The chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e showed a high degree of homology compared to other reported chloroplast genomes of Anacardiaceae species, including \u003cem\u003eDracontomelon delavayi\u003c/em\u003e, \u003cem\u003eLannea coromandelica\u003c/em\u003e, \u003cem\u003ePistacia nitida\u003c/em\u003e, \u003cem\u003eSpondias paniculata\u003c/em\u003e, and \u003cem\u003eSwintonia reticulata\u003c/em\u003e. The genome lengths of these species ranged from 159,485 to 162,509 bp, with a maximum difference of 3,024 bp. The overall GC content is comparable to that of other species in the Anacardiaceae family, such as \u003cem\u003eRhus chinensis\u003c/em\u003e (37.79%), \u003cem\u003ePistacia weinmannifolia\u003c/em\u003e (37.84%), \u003cem\u003eToxicodendron vernicifluum\u003c/em\u003e (37.96%), and \u003cem\u003eCotinus species\u003c/em\u003e (37.9%-38.1%)\u003c/p\u003e \u003cp\u003e(Wang et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Zheng et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, Liu et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). The number and types of genes were also very similar, reflecting the highly conserved characteristics of chloroplast genomes. The homology among closely related woody species is typically considered reasonable due to their long generation times and the relatively low number of substitutions occurring over a given period. Substitutions from parent plants can only be passed to offspring during the process of germination.\u003c/p\u003e \u003cp\u003eThe study of codon preference aids in understanding the evolutionary processes of plant species and optimizing the expression of exogenous genes in chloroplasts, enabling the prediction of gene function and expression levels (Li et al., \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2019b\u003c/span\u003e). Consistent with previous findings, \u003cem\u003eBuchanania latifolia\u003c/em\u003e demonstrates a preference for A or U bases in its chloroplast genome codons, a common trait in plant chloroplast genomes (Zhou et al., \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). In B. latifolia, 30 high-frequency codons were identified, with 29 of them ending with an A or U base, likely influenced by natural selection and mutations (Necşulea and Lobry, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Previous studies have also noted that in the chloroplast genomes of Anacardiaceae, high-frequency codons tend to utilize A or U bases as the third codon base (Liu et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, Xin et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, Wang et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe most prevalent SSRs in the chloroplast genomes of \u003cem\u003eB. latifolia\u003c/em\u003e were mononucleotide repeats. Similar to other plants, chloroplast SSRs predominantly consist of short poly-A or poly-T repeats, with mononucleotide repeats being the most common forms (Tao et al., \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, Vu et al., \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Djedid et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2021\u003c/span\u003e, Provan et al., \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2001\u003c/span\u003e, Yang et al., \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Moreover, the majority of SSRs are located in the LSC and SSC regions, consistent with previous findings on chloroplast genomes (Alshegaihi, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2024\u003c/span\u003e, Liu et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, Wang et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Palindrome sequences accounted for 39.68%, forward repeated sequences for 57.14%, and reverse sequences for 3.17%. No complementary sequences were found. Forward and palindromic repeats were the most common repeat types, with most dispersed repeats being less than 40 bp, as reported in previous studies (Liang et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Kirov et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Tian et al., \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2021\u003c/span\u003e, Yuan et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2005\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe IR region of the chloroplast genome is thought to be the most conservative section. The expansion and contraction of the IR region are pivotal factors influencing the length variation observed in plant chloroplast genomes, typically categorized into two types (Yi et al., \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2013\u003c/span\u003e, Zhang et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Such expansions and contractions in the IR region across most species manifest as minor deviations in the IR/SC boundary within a few fixed genes, which may lead to pseudogenization of certain genes. The examination of the chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e and the Anacardiaceae family in this study aligns with prior reports in angiosperms (Liu et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, Wang et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Xin et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Typically, the LSC/IRb boundary is situated on or near \u003cem\u003erps19\u003c/em\u003e, \u003cem\u003erpl2\u003c/em\u003e, or \u003cem\u003erpl22\u003c/em\u003e, while IRb/SSC is generally positioned on \u003cem\u003eycf1\u003c/em\u003e or between \u003cem\u003eycf1\u003c/em\u003e and \u003cem\u003endhF\u003c/em\u003e. The SSC/IRa boundary typically lies on \u003cem\u003eycf1\u003c/em\u003e, and the IRa/LSC boundary usually falls on or near \u003cem\u003erps19\u003c/em\u003e, \u003cem\u003erpl2\u003c/em\u003e, \u003cem\u003erpl12\u003c/em\u003e, and \u003cem\u003etrnH\u003c/em\u003e (Alshegaihi, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIntergenic spacers are more divergent than introns and protein-coding sequences (Meng et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, pseudogenes suffer the same fate as intergenic spacers due to a lack of functional importance, leading to less conservative strains. Pseudogenization was common in the evolution of chloroplast genomes, such as \u003cem\u003eaccD\u003c/em\u003e, \u003cem\u003eccsA\u003c/em\u003e, \u003cem\u003eycf1\u003c/em\u003e, \u003cem\u003erps19\u003c/em\u003e and \u003cem\u003epsbB\u003c/em\u003e pseudogenes (Krawczyk et al., \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, Li et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The most divergent genes among the six Anacardiaceae species were two pseudogenes \u003cem\u003eaccD\u003c/em\u003e and \u003cem\u003eycf1\u003c/em\u003e, an intron of the \u003cem\u003erps16\u003c/em\u003e genes, and the intergenic spacer \u003cem\u003endhF-rpl32\u003c/em\u003e. These genes or regions can be utilized for developing molecular markers for species identification and population studies (Magdy et al., \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).,\u003c/p\u003e \u003cp\u003eThe synonymous (Ks) and non-synonymous (Ka) nucleotide substitution pattern is a well-recognized marker for assessing genome evolution; and the Ka/Ks ratio reflects selection pressure on genes (Yang and Nielsen, \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2000\u003c/span\u003e, Guo et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Ka/Ks\u0026thinsp;\u0026lt;\u0026thinsp;1, Ka/Ks\u0026thinsp;=\u0026thinsp;1, and Ka/Ks\u0026thinsp;\u0026gt;\u0026thinsp;1 indicate genes that underwent purifying, neutral, and positive selections, respectively (Yang and Nielsen, \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Generally, synonymous mutations occur more frequently than nonsynonymous mutations within genes, causing the Ka/Ks values to be below 1 (Makałowski and Boguski, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e1998\u003c/span\u003e). Our results using \u003cem\u003eB. latifolia\u003c/em\u003e and five species from the Anacardiaceae family indicate that only the \u003cem\u003epsbT\u003c/em\u003e gene in \u003cem\u003eL. coromandelica\u003c/em\u003e and the \u003cem\u003erpl22\u003c/em\u003e, \u003cem\u003erpl32\u003c/em\u003e, \u003cem\u003erps16\u003c/em\u003e and \u003cem\u003eycf2\u003c/em\u003e genes in \u003cem\u003eS. paniculata\u003c/em\u003e have Ka/Ks values\u0026thinsp;\u0026gt;\u0026thinsp;1, suggesting strong positive selection acting on these two genes. Ka/Ks values for all other detected genes are below 1, indicating widespread purifying selection on these chloroplast genomes.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003eApplication on Phylogenetic Reconstruction\u003c/h2\u003e \u003cp\u003eThe informative sites provided by single genes or gene combinations are often limited. Differences in the evolutionary trajectories of various gene sequences can result in low resolution of the constructed phylogenetic tree, posing challenges in elucidating the evolutionary relationships within the Anacardiaceae family. To optimize phylogenetic outcomes, we conducted a phylogenomic analysis of Anacardiaceae based on complete chloroplast genomes using the maximum likelihood method. Overall, branch support at the generic level was generally high, with only relatively lower support observed for the branch of the genus Rhus and its sister groups (BS\u0026thinsp;=\u0026thinsp;68), while support for all other branches was robust (BS\u0026thinsp;=\u0026thinsp;100). This suggests that our analysis accurately reflects the phylogenetic relationships within Anacardiaceae. \u003cem\u003eB. latifolia\u003c/em\u003e is sister to \u003cem\u003eChoerospondias\u003c/em\u003e, \u003cem\u003eLannea\u003c/em\u003e, and \u003cem\u003eSclerocarya\u003c/em\u003e, consistent with previous phylogenetic trees constructed using sequences such as nuclear ribosomal external transcribed spacer (ETS), the chloroplast \u003cem\u003etrnL\u003c/em\u003e intron and \u003cem\u003etrnL-F\u003c/em\u003e intergenic spacer (\u003cem\u003etrnL-F\u003c/em\u003e region), and the chloroplast rps16 intron (Weeks et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). However, \u003cem\u003eChoerospondias\u003c/em\u003e, \u003cem\u003eLannea\u003c/em\u003e, and \u003cem\u003eSclerocarya\u003c/em\u003e all belong to the tribe/subfamily Spondiadeae/Spondioideae. Phylogenetically, they are distant from \u003cem\u003eAnacardium\u003c/em\u003e and \u003cem\u003eMangifera\u003c/em\u003e, which are in the same tribe/subfamily Anacardieae/Anacardioideae as \u003cem\u003eBuchanania\u003c/em\u003e. Although \u003cem\u003eBuchanania\u003c/em\u003e is typically considered a member of the tribe/subfamily Anacardieae/Anacardioideae, it is here regarded as a sister group to those traditionally belonging to the tribe/subfamily Spondiadeae/Spondioideae, which aligns with Weeks' view (Weeks et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2014\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e"},{"header":"Conclusions","content":"\u003cp\u003eIn this study, the complete chloroplast genome sequence of \u003cem\u003eB. latifolia\u003c/em\u003e was assembled and annotated de novo. The chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e exhibits a typical quadripartite structure, with a length of 160,088 bp, containing 88 protein-coding sequences (CDS), 37 tRNA genes, and 8 rRNA genes, with a GC content of 37.7%. A total of 99 SSR loci and 63 repeat sequences were identified, which can be utilized for marker development, phylogenetic and population studies of \u003cem\u003eB. latifolia\u003c/em\u003e. Codon usage analysis revealed a preference for Leu codons ending with A/U. Additionally, the study investigated IR boundaries, DNA polymorphism, positive selection suites, and phylogenetic position. Comparative analysis with five other species from the Anacardiaceae family confirmed the nearly identical and highly conserved chloroplast genome features of \u003cem\u003eB. latifolia\u003c/em\u003e, which can be valuable for understanding the plastid evolution and evolutionary relationships within Anacardiaceae. Phylogenetic analysis reveals that \u003cem\u003eB. latifolia\u003c/em\u003e is positioned at the base of Anacardiaceae, sister to \u003cem\u003eChoerospondias\u003c/em\u003e, \u003cem\u003eLannea\u003c/em\u003e, and \u003cem\u003eSclerocarya\u003c/em\u003e. These fndings could provide important genetic information for further research into breeding of Anacardiaceae, phylogeny, and evolution of \u003cem\u003eB. latifolia\u003c/em\u003e. This study reveals the structural characteristics of the chloroplast genome, SSR loci, and phylogenetic analysis, providing valuable genetic information for understanding the origin and evolution of Anacardiaceae in the future.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003eData Availability\u003c/p\u003e\n\u003cp\u003eThe genome sequence data that support the fndings of this study are openly available in GenBank of NCBI at https://www.ncbi.nlm.nih.gov/ under the accession OM000214.1. The associated BioProject, SRA, and Bio-Sample numbers are PRJNA783515, SRR17036892, and SAMN23436132 respectively.\u003c/p\u003e\n\u003cp\u003eFunding\u003c/p\u003e\n\u003cp\u003eThis research was funded by the National Natural Science Foundation of China (grant No. 42161015). The Key Project of Basic Research of Yunnan Province, China (202202AS07337) to Gao Chen. This work is supported by Yunnan Science and Technology Talent and platform Program (202305AM340008).\u003c/p\u003e\n\u003cp\u003eAcknowledgements\u003c/p\u003e\n\u003cp\u003eThanks to Zhefei Zeng and Kaiwen Jiang for their support of field sampling.\u003c/p\u003e\n\u003cp\u003eAuthors and Affiliations\u003c/p\u003e\n\u003cp\u003e(1) Yunnan Key Laboratory of Plateau Wetland Conservation, Restoration and Ecological Services; National Plateau Wetlands Research Center; Dianchi Lake Ecosystem Observation and Research Station of Yunnan Province; Southwest Forestry University, Kunming, PR, 650224, China\u003c/p\u003e\n\u003cp\u003eChunmin Mao, Rui Rao, WanTing Chen, Liangliang Yue\u003c/p\u003e\n\u003cp\u003e(2) Technology Centre of Kunming Customs\u003c/p\u003e\n\u003cp\u003eQuwen Lei\u003c/p\u003e\n\u003cp\u003eContributions\u003c/p\u003e\n\u003cp\u003eLLY designed the study. CMM performed statistical and bioinformatic analyzes and wrote the first draft. RR proofreads the image text and provides suggestions for revision. QWL sampling, data analysis. WTC Molecular Materials Management. CMM and RR contribute equally in this study. All-authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003eCorresponding author\u003c/p\u003e\n\u003cp\u003eCorrespondence to Liangliang Yue([email protected])\u003c/p\u003e\n\u003cp\u003eEthics declarations\u003c/p\u003e\n\u003cp\u003eEthical approval\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003eInformed consent.\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003eConflict of interest\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003eAdditional information\u003c/p\u003e\n\u003cp\u003eCommunicated by Ewa Liangliang Yue.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eAlshegaihi R M 2024. The complete chloroplast genome of the halophyte flowering plant Suaeda monoica from Jeddah, Saudi Arabia. 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Forestry Studies in China [J], 10: 235-242. https://doi.org/10.1007/s11632-008-0047-1\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[{"identity":"55ee872e-f1dd-43af-92af-b2fe6d8f06fa","identifier":"10.13039/501100001809","name":"National Natural Science Foundation of China","awardNumber":"42161015","order_by":0}],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Southwest Forestry University","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":"Buchanania latifolia Roxb., Anacardiaceae, Chloroplast genome, SSR loci, Phylogeny","lastPublishedDoi":"10.21203/rs.3.rs-4552236/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4552236/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe chloroplast (cp) genomes are valuable resource with multiple applications, encompassing species identification, phylogenetic reconstruction, and evolutionary investigations. In this study, the complete chloroplast genome sequence of \u003cem\u003eBuchanania latifolia\u003c/em\u003ewas de novo sequenced, assembled and annotated. The chloroplast genome of \u003cem\u003eB. latifolia\u003c/em\u003e exhibits a typical quadripartite structure, with a total length of 160,088 bp, containing 88 protein-coding sequences (CDS), 37 tRNA genes, and 8 rRNA genes, with an overall GC content of 37.7%. A total of 99 SSR loci and 63 repeat sequences were identified, which can be utilized for marker development, phylogenetic and population studies of \u003cem\u003eB. latifolia\u003c/em\u003e. Codon usage analysis revealed a preference for Leu codons ending with A/U. Additionally, the study investigated IR boundaries, DNA polymorphism, positive selection suites, and phylogenetic position. Comparative analysis with five other species from the Anacardiaceae family confirmed the nearly identical and highly conserved chloroplast genome features of \u003cem\u003eB. latifolia\u003c/em\u003e, which can be valuable for understanding the plastid evolution and evolutionary relationships within Anacardiaceae. Phylogenetic analysis reveals that \u003cem\u003eB. latifolia\u003c/em\u003eis positioned at the base of Anacardiaceae, sister to\u003cem\u003e Choerospondias axillaris\u003c/em\u003e,\u003cem\u003e Lannea coromandelica\u003c/em\u003e, and \u003cem\u003eSclerocarya birrea\u003c/em\u003e. These findings could provide important genetic information for further research into breeding of Anacardiaceae, phylogeny, and evolution of \u003cem\u003eB. latifolia\u003c/em\u003e.\u003c/p\u003e","manuscriptTitle":"Complete chloroplast genome sequence of Buchanania latifolia (Anacardiaceae): genome structure, and phylogenetic relationships","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-06-11 11:16:09","doi":"10.21203/rs.3.rs-4552236/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":"3b3ad84d-b579-4246-8a25-7e096ad14931","owner":[],"postedDate":"June 11th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":32997943,"name":"Molecular Genetics"},{"id":32997944,"name":"Plant Molecular Biology and Genetics"}],"tags":[],"updatedAt":"2024-06-11T11:16:09+00:00","versionOfRecord":[],"versionCreatedAt":"2024-06-11 11:16:09","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4552236","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4552236","identity":"rs-4552236","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}