Comparative Analysis of Chloroplast Genome Characteristics among Three Cultivated Types of Chrysanthemum Varieties

doi:10.13560/j.cnki.biotech.bull.1985.2025-1432

Abstract

Abstract:

Objective To explore the chloroplast genome characteristics and phylogenetic relationships of Chrysanthemum cultivars of different cultivation types, providing a theoretical basis for research on the phylogeny, species identification and germplasm conservation of the genus Chrysanthemum. Method Chloroplast genome sequencing, assembly and annotation were performed on the groundcover Chrysanthemum × morifolium ‘Bei Lin Qiu Yun’, the single-headed cut flower C. morifolium ‘Sheng Xue’, and the traditional large-flowered C. morifolium ‘Fen Shi Ba’. Analysis of genomic structure, simple sequence repeats (SSRs), codon usage bias, inverted repeat (IR) boundary regions and collinearity characteristics was conducted. For phylogenetic analysis, nine chloroplast genome sequences, including eight from Chrysanthemum species and one outgroup from the Asteraceae family, were downloaded from GenBank. Result The chloroplast genomes of the three cultivars all exhibited a typical double-stranded circular quadripartite structure, ranging in length from 151 060 to 151 073 bp, with a GC content of 37.5%. A total of 131 genes were annotated in each genome, including 86 protein-coding genes, 37 transfer RNA (tRNA) genes, and 8 ribosomal RNA (rRNA) genes. Whole-genome alignment revealed high overall conservation of the chloroplast genomes: the IR regions showed higher conservation than the single-copy (SC) regions, and the non-coding regions had a higher mutation frequency than the coding regions, with no large-scale gene rearrangements detected. Boundary gene types were consistent across all samples. C. morifolium ‘Sheng Xue’ and C. morifolium ‘Fen Shi Ba’ each contained 68 SSRs, whereas C. morifolium ‘Bei Lin Qiu Yun’ contained 70 SSRs. Mononucleotide repeats (A/T-type) dominated the SSRs in all three cultivars, with no hexanucleotide repeats detected. Codon usage bias was weak (CAI: 0.168‒0.169; ENC: 49.127‒49.420), with a preference for A/U-ending codons. Selection pressure was identified as the key factor influencing codon usage bias. Phylogenetic analysis showed that C. morifolium ‘Sheng Xue’ and C. morifolium ‘Fen Shi Ba’ were closely related to cultivated C. morifolium, while C. morifolium ‘Bei Lin Qiu Yun’ was closely related to wild Chrysanthemum species. Conclusion The three Chrysanthemum cultivars of different cultivation types have a conserved chloroplast genomic structure and gene content. The groundcover C. morifolium is more closely related to wild species, whereas the traditional large-flowered C. morifolium and single-headed cut C. morifolium are more closely related to cultivated varieties.

Key words: Chrysanthemum, Chrysanthemum × morifolium, chloroplast genome, phylogenetic analysis, sequence characteristics, codon usage bias, SSR

LIU Rou-zhi, WU Jia-qi, WU Hui-ye, YUAN Jing-yi, CHENG Tang-ren, ZHANG Qi-xiang, YUAN Cun-quan. Comparative Analysis of Chloroplast Genome Characteristics among Three Cultivated Types of Chrysanthemum Varieties[J]. Biotechnology Bulletin, 2026, 42(8): 239-250.

Figures/Tables 12

References 55

[1]	Yuan HW, Jiang S, Liu YK, et al. The flower head of Chrysanthemum morifolium Ramat. (Juhua): a paradigm of flowers serving as Chinese dietary herbal medicine [J]. J Ethnopharmacol, 2020, 261: 113043.
[2]	Hu J, Ma W, Li N, et al. Antioxidant and anti-inflammatory flavonoids from the flowers of chuju, a medical cultivar of Chrysanthemum morifolim Ramat [J]. J Mex Chem Soc, 2018, 61(4): 282-289.
[3]	Pan B, Du YX, Chen QB, et al. China’s Chrysanthemum in the global market: evaluating the international competitiveness and influencing factors [J]. Front Sustain Food Syst, 2025, 9: 1521709.
[4]	Van Huylenbroeck J. Ornamental Crops [M]. Cham: Springer International Publishing, 2018.
[5]	李鸿渐, 邵健文. 中国菊花品种资源的调查收集与分类 [J]. 南京农业大学学报, 1990, 13(1): 30-36.
	Li HJ, Shao JW. Investigation, collection and classification of Chrysanthemum cultivars in China [J]. J Nanjing Agric Univ, 1990, 13(1): 30-36.
[6]	Zhang Y, Luo XY, Zhu J, et al. A classification study for chrysanthemum (Chrysanthemum × grandiflorum Tzvelv.) cultivars based on multivariate statistical analyses [J]. J Sytematics Evolution, 2014, 52(5): 612-628.
[7]	Yang YY, Wen C, Ma N, et al. Heterosis and genetic analysis of branching in cut-flower chrysanthemums [J]. Euphytica, 2015, 205(3): 915-925.
[8]	韩勇. 切花菊品质综合评价体系的构建及性状遗传分析 [D]. 南京: 南京农业大学, 2011.
	Han Y. Study on comprehensive evaluation system of cut chrysanthemum and genetic analysis of main characters [D]. Nanjing: Nanjing Agricultural University, 2011.
[9]	Sun MX, Zhang JY, Lu JS, et al. Intelligent evaluation of ornamental characteristics of garden chrysanthemums using unmanned aerial vehicle (UAV)-based RGB imagery [J]. Sci Hortic, 2025, 350: 114374.
[10]	Kirchhoff H. Chloroplast ultrastructure in plants [J]. New Phytol, 2019, 223(2): 565-574.
[11]	Li MM, Geng ML, Zhou YF, et al. Integrative chloroplast genomics unravels phylogenetic discordance in Glehnia (Apiaceae) [J]. Ind Crops Prod, 2025, 236: 121978.
[12]	Zhao CH, Li XY, Lan X, et al. Cytotype classification and genetic diversity of Platostoma palustre revealed by rDNA localization and chloroplast genome [J]. BMC Genom, 2025, 26: 937.
[13]	Yin DM, Wang Y, Zhang XG, et al. Development of chloroplast genome resources for peanut (Arachis hypogaea L.) and other species of Arachis [J]. Sci Rep, 2017, 7: 11649.
[14]	Zhu AD, Guo WH, Gupta S, et al. Evolutionary dynamics of the plastid inverted repeat: the effects of expansion, contraction, and loss on substitution rates [J]. New Phytol, 2016, 209(4): 1747-1756.
[15]	Park SK, Kwon SJ, Park J, et al. Chloroplast genome of white wild Chrysanthemum, Dendranthema sp. K247003, as genetic barcode [J]. J Species Res, 2015, 4(2): 152-158.
[16]	Tyagi S, Jung JA, Kim JS, et al. The complete chloroplast genome of an economic plant, Chrysanthemum morifolium ‘Baekma’ [J]. Mitochondrial DNA Part B, 2019, 4(2): 3133-3134.
[17]	Zeng T, Li JW, Fu Q, et al. The complete chloroplast of Chrysanthemum × morifolium ‘fubaiju’ [J]. Mitochondrial DNA Part B, 2021, 6(10): 3062-3063.
[18]	Xu YF, Liao BR, Ostevik KL, et al. The maternal donor of Chrysanthemum cultivars revealed by comparative analysis of the chloroplast genome [J]. Front Plant Sci, 2022, 13: 923442.
[19]	Parvathy ST, Udayasuriyan V, Bhadana V. Codon usage bias [J]. Mol Biol Rep, 2022, 49(1): 539-565.
[20]	刘慧, 王梦醒, 岳文杰, 等. 糜子叶绿体基因组密码子使用偏性的分析 [J]. 植物科学学报, 2017, 35(3): 362-371.
	Liu H, Wang MX, Yue WJ, et al. Analysis of codon usage in the chloroplast genome of broomcorn millet (Panicum miliaceum L.) [J]. Plant Sci J, 2017, 35(3): 362-371.
[21]	Jin JJ, Yu WB, Yang JB, et al. GetOrganelle: a fast and versatile toolkit for accurate de novo assembly of organelle genomes [J]. Genome Biol, 2020, 21: 241.
[22]	Liu C, Shi LC, Zhu YJ, et al. CpGAVAS, an integrated web server for the annotation, visualization, analysis, and GenBank submission of completely sequenced chloroplast genome sequences [J]. BMC Genom, 2012, 13(1): 715.
[23]	Kearse M, Moir R, Wilson A, et al. Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data [J]. Bioinformatics, 2012, 28(12): 1647-1649.
[24]	Greiner S, Lehwark P, Bock R. OrganellarGenomeDRAW (OGDRAW) version 1.3.1: expanded toolkit for the graphical visualization of organellar genomes [J]. Nucleic Acids Res, 2019, 47(W1): W59-W64.
[25]	Li HE, Guo QQ, Xu L, et al. CPJSdraw: analysis and visualization of junction sites of chloroplast genomes [J]. PeerJ, 2023, 11: e15326.
[26]	Frazer KA, Pachter L, Poliakov A, et al. VISTA: computational tools for comparative genomics [J]. Nucleic Acids Res, 2004, 32(): W273-W279.
[27]	Darling ACE, Mau B, Blattner FR, et al. Mauve: multiple alignment of conserved genomic sequence with rearrangements [J]. Genome Res, 2004, 14(7): 1394-1403.
[28]	Thiel T, Michalek W, Varshney R, et al. Exploiting EST databases for the development and characterization of gene-derived SSR-markers in barley (Hordeum vulgare L.) [J]. Theor Appl Genet, 2003, 106(3): 411-422.
[29]	Wang ZJ, Cai QW, Wang Y, et al. Comparative analysis of codon bias in the chloroplast genomes of Theaceae species [J]. Front Genet, 2022, 13: 824610.
[30]	Sharp PM, Li WH. An evolutionary perspective on synonymous codon usage in unicellular organisms [J]. J Mol Evol, 1986, 24(1/2): 28-38.
[31]	尚明照, 刘方, 华金平, 等. 陆地棉叶绿体基因组密码子使用偏性的分析 [J]. 中国农业科学, 2011, 44(2): 245-253.
	Shang MZ, Liu F, Hua JP, et al. Analysis on codon usage of chloroplast genome of Gossypium hirsutum [J]. Sci Agric Sin, 2011, 44(2): 245-253.
[32]	秦政, 郑永杰, 桂丽静, 等. 樟树叶绿体基因组密码子偏好性分析 [J]. 广西植物, 2018, 38(10): 1346-1355.
	Qin Z, Zheng YJ, Gui LJ, et al. Codon usage bias analysis of chloroplast genome of Camphora tree (Cinnamomum camphora) [J]. Guihaia, 2018, 38(10): 1346-1355.
[33]	Fu LF, Xin ZB, Wen F, et al. Complete chloroplast genome sequence of Elatostema dissectum (Urticaceae) [J]. Mitochondrial DNA Part B, 2019, 4(1): 838-839.
[34]	Zhang D, Gao FL, Jakovlić I, et al. PhyloSuite: an integrated and scalable desktop platform for streamlined molecular sequence data management and evolutionary phylogenetics studies [J]. Mol Ecol Resour, 2020, 20(1): 348-355.
[35]	Ronquist F, Teslenko M, van der Mark P, et al. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space [J]. Syst Biol, 2012, 61(3): 539-542.
[36]	Suchard MA, Lemey P, Baele G, et al. Bayesian phylogenetic and phylodynamic data integration using BEAST 1.10 [J]. Virus Evol, 2018, 4(1): vey016.
[37]	Khade YP, Mainkar P, Chandanshive A, et al. Harnessing chloroplast SSRs to decipher genetic diversity in underutilized Allium species [J]. Front Plant Sci, 2025, 16: 1645145.
[38]	Xia Y, Hu ZG, Li XW, et al. The complete chloroplast genome sequence of Chrysanthemum indicum [J]. Mitochondrial DNA Part A, 2016, 27(6): 4668-4669.
[39]	Baek J, Park S, Lee J, et al. The complete chloroplast genome of Chrysanthemum zawadskii Herbich (Asteraceae) isolated in Korea [J]. Mitochondrial DNA Part B, 2021, 6(7): 1956-1958.
[40]	Guo YY, Yang JX, Li HK, et al. Chloroplast genomes of two species of Cypripedium: expanded genome size and proliferation of AT-biased repeat sequences [J]. Front Plant Sci, 2021, 12: 609729.
[41]	Ma YP, Chen MM, Wei JX, et al. Origin of Chrysanthemum cultivars—Evidence from nuclear low-copy LFY gene sequences [J]. Biochem Syst Ecol, 2016, 65: 129-136.
[42]	Yang Z, Ma WX, Yang XH, et al. Plastome phylogenomics provide new perspective into the phylogeny and evolution of Betulaceae (Fagales) [J]. BMC Plant Biol, 2022, 22: 611.
[43]	Meng XX, Xian YF, Xiang L, et al. Complete chloroplast genomes from Sanguisorba: identity and variation among four species [J]. Molecules, 2018, 23(9): 2137.
[44]	Zhang L, Zhang CP, Zhang ED, et al. Hybridization of Lycium qingshuiheense and L. ningxiaense revealed using morphological, SSR markers, and chloroplast genomic data [J]. BMC Genom, 2025, 26: 965.
[45]	Gong YH, Qin YL, Luo T, et al. Complete chloroplast genome sequence and phylogenetic analysis of Lactuca sativa var. ramosa hort [J]. Genet Resour Crop Evol, 2025, 72(6): 7531-7546.
[46]	Liu X, Zhou BY, Yang HY, et al. Sequencing and analysis of Chrysanthemum carinatum schousb and Kalimeris indica. The complete chloroplast genomes reveal two inversions and rbcL as barcoding of the vegetable [J]. Molecules, 2018, 23(6): 1358.
[47]	Wu CW, Liu R, Li CH, et al. Comparative analysis of codon usage bias and phylogenetic relationships in chloroplast genomes across 49 Dendrobium species [J]. BMC Plant Biol, 2025, 25: 1411.
[48]	Xie P, Wu JR, Lu MY, et al. Assembly and comparative analysis of the complete mitochondrial genome of Fritillaria ussuriensis Maxim. (Liliales: Liliaceae), an endangered medicinal plant [J]. BMC Genom, 2024, 25: 773.
[49]	Yengkhom S, Uddin A, Chakraborty S. Deciphering codon usage patterns and evolutionary forces in chloroplast genes of Camellia sinensis var. assamica and Camellia sinensis var. sinensis in comparison to Camellia pubicosta [J]. J Integr Agric, 2019, 18(12): 2771-2785.
[50]	Li RX, Wang B, Xiao SQ, et al. Characterization of the complete chloroplast genome and comparative analysis of the phylogeny and codon usage bias of three Yunnan wild rice species [J]. Front Plant Sci, 2025, 16: 1555104.
[51]	Yang R, Xue Y, He XF, et al. Comprehensive analysis of chloroplast genomes in leguminous forage species: codon usage, phylogenetic relationships, and evolutionary insights [J]. Agronomy, 2025, 15(4): 765.
[52]	Shi NX, Yuan YW, Huang RJ, et al. Analysis of codon usage patterns in complete plastomes of four medicinal Polygonatum species (Asparagaceae) [J]. Front Genet, 2024, 15: 1401013.
[53]	Yao ZL, Miao JS, Xu WD, et al. The complete chloroplast genome sequence of a cultivar of Chrysanthemum, Chrysanthemum × morifolium ‘Hangbaiju’ (Asteraceae) [J]. Mitochondrial DNA Part B, 2024, 9(3): 403-407.
[54]	Duan YY, Wang YQ, Ding WY, et al. Comparative and phylogenetic analysis of the chloroplast genomes of four commonly used medicinal cultivars of Chrysanthemums morifolium [J]. BMC Plant Biol, 2024, 24: 992.
[55]	Chong XR, Zhang F, Wu YY, et al. A SNP-enabled assessment of genetic diversity, evolutionary relationships and the identification of candidate genes in Chrysanthemum [J]. Genome Biol Evol, 2016, 8(12): 3661-3671.

功能 Functions	基因分组 Gene grouping	基因名 Gene name
自我复制 Self-replication	核糖体大亚基 Large ribosomal subunit	rpl32， rpl23（2）， rpl2（2）， rpl22， rpl16， rpl14， rpl36， rpl20， rpl33*
自我复制 Self-replication	核糖体小亚基 Small ribosomal subunit	rps15， rps12， rps12-5end， rps12-3end， rps7（2）， rps19， rps3， rps8， rps11， rps18， rps4， rps14， rps2， rps16*
	RNA聚合酶 RNA polymerase	rpoA， rpoC2， rpoC1， rpoB
	核糖体RNA Ribosomal RNA	rrn5（2）， rrn4.5（2）， rrn23（2）， rrn16（2）
	转运RNA Transfer RNA	trnL-UAG， trnN-GUU（2）， trnR-ACG（2）， trnA-UGC（2）， trnI-GAU（2）， trnI-CAU（2）， trnV-GAC（2）， trnL-CAA（2）， trnM-CAU（2）， trnP-UGG， trnW-CCA， trnV-UAC， trnF-GAA， trnL-UAA， trnT-UGU， trnS-GGA， trnG-GCC， trnS-UGA， trnT-GGU， trnG-UCC， trnR-UCU， trnE-UUC， trnY-GUA， trnD-GUC， trnC-GCA， trnS-GCU， trnQ-UUG， trnK-UUU， trnH-GUG
光合作用 Photosynthesis	光系统I基因 Photosystem I gene	psaC， psaJ， psaI， psaA， psaB
	光系统II基因 Photosystem II gene	psbH，，psbT， psbB， psbE， psbF， psbL， psbJ， psbZ， psbC， psbD， psbM， psbI， psbK， psbA
	NADH氧化还原酶 NADH dehydrogenase	ndhH， ndhA， ndhI， ndhG， ndhE， ndhD， ndhF， ndhB（2）， ndhC， ndhK， ndhJ*
	细胞色素复合物基因 Cytochrome complex gene	petB， petG， petL， petA， petN*
	ATP合酶 ATP synthase	atpB， atpE， atpA， atpF， atpH， atpI*
	Rubisco大亚基 Large subunit of Rubisco	rbcL
其他基因 Other gene	成熟酶 Maturase	matK
其他基因 Other gene	翻译起始因子 Translational initiation factor	infA
	蛋白酶 Protease	clpP1**
	被膜蛋白 Envelop membrane protein	cemA
	乙酰辅酶A亚基 Acetyl-CoA carboxylase subunit	accD
	C型细胞色素合成基因 C-type cytochrome synthesis gene	ccsA
功能未知基因Genes of unknown function	保守的假定叶绿体开放阅读框ORF Conserved hypothetical chloroplast ORF	ycf1， φycf1， ycf15（2）， ycf2（2）， ycf3， ycf4

功能 Functions	基因分组 Gene grouping	基因名 Gene name
自我复制 Self-replication	核糖体大亚基 Large ribosomal subunit	rpl32， rpl23（2）， rpl2（2）， rpl22， rpl16， rpl14， rpl36， rpl20， rpl33*
自我复制 Self-replication	核糖体小亚基 Small ribosomal subunit	rps15， rps12， rps12-5end， rps12-3end， rps7（2）， rps19， rps3， rps8， rps11， rps18， rps4， rps14， rps2， rps16*
	RNA聚合酶 RNA polymerase	rpoA， rpoC2， rpoC1， rpoB
	核糖体RNA Ribosomal RNA	rrn5（2）， rrn4.5（2）， rrn23（2）， rrn16（2）
	转运RNA Transfer RNA	trnL-UAG， trnN-GUU（2）， trnR-ACG（2）， trnA-UGC（2）， trnI-GAU（2）， trnI-CAU（2）， trnV-GAC（2）， trnL-CAA（2）， trnM-CAU（2）， trnP-UGG， trnW-CCA， trnV-UAC， trnF-GAA， trnL-UAA， trnT-UGU， trnS-GGA， trnG-GCC， trnS-UGA， trnT-GGU， trnG-UCC， trnR-UCU， trnE-UUC， trnY-GUA， trnD-GUC， trnC-GCA， trnS-GCU， trnQ-UUG， trnK-UUU， trnH-GUG
光合作用 Photosynthesis	光系统I基因 Photosystem I gene	psaC， psaJ， psaI， psaA， psaB
	光系统II基因 Photosystem II gene	psbH，，psbT， psbB， psbE， psbF， psbL， psbJ， psbZ， psbC， psbD， psbM， psbI， psbK， psbA
	NADH氧化还原酶 NADH dehydrogenase	ndhH， ndhA， ndhI， ndhG， ndhE， ndhD， ndhF， ndhB（2）， ndhC， ndhK， ndhJ*
	细胞色素复合物基因 Cytochrome complex gene	petB， petG， petL， petA， petN*
	ATP合酶 ATP synthase	atpB， atpE， atpA， atpF， atpH， atpI*
	Rubisco大亚基 Large subunit of Rubisco	rbcL
其他基因 Other gene	成熟酶 Maturase	matK
其他基因 Other gene	翻译起始因子 Translational initiation factor	infA
	蛋白酶 Protease	clpP1**
	被膜蛋白 Envelop membrane protein	cemA
	乙酰辅酶A亚基 Acetyl-CoA carboxylase subunit	accD
	C型细胞色素合成基因 C-type cytochrome synthesis gene	ccsA
功能未知基因Genes of unknown function	保守的假定叶绿体开放阅读框ORF Conserved hypothetical chloroplast ORF	ycf1， φycf1， ycf15（2）， ycf2（2）， ycf3， ycf4

品种名 Cultivar	密码子适应指数CAI	密码子偏性指数CBI	有效密码子数ENC	GC1 （%）	GC2 （%）	GC3 （%）
C. morifolium ‘Bei Lin Qiu Yun’	0.168	-0.107	49.127	45.95	38.05	29.08
C. morifolium ‘Sheng Xue’	0.169	-0.102	49.420	46.64	38.70	29.46
C. morifolium ‘Fen Shi Ba’	0.169	-0.103	49.407	46.89	38.88	29.44

品种名 Cultivar	密码子适应指数CAI	密码子偏性指数CBI	有效密码子数ENC	GC1 （%）	GC2 （%）	GC3 （%）
C. morifolium ‘Bei Lin Qiu Yun’	0.168	-0.107	49.127	45.95	38.05	29.08
C. morifolium ‘Sheng Xue’	0.169	-0.102	49.420	46.64	38.70	29.46
C. morifolium ‘Fen Shi Ba’	0.169	-0.103	49.407	46.89	38.88	29.44