Crops ›› 2023, Vol. 39 ›› Issue (1): 30-37.doi: 10.16035/j.issn.1001-7283.2023.01.005

Previous Articles     Next Articles

Genetic Diversity Analysis of Sophora flavescens Ait. Germplasm Resources Based on cpSSR Markers

Song Yun1,2(), Zhang Xinrui1, He Jiaxin1, Li Zheng1, Sun Zhe1, Li Aoxuan1, Qiao Yonggang1,2()   

  1. 1College of Life Sciences, Shanxi Agricultural University, Jinzhong 030801, Shanxi, China
    2Shanxi Key Laboratory for Modernization of TCVM, Jinzhong 030801, Shanxi, China
  • Received:2021-08-23 Revised:2021-10-22 Online:2023-02-15 Published:2023-02-22

RichHTML

8

PDF (PC)

184

Abstract:

A total of 150 Sophora flavescens plants from 15 different habitats were used as materials, and cpSSR molecular marker technology was used to explore the genetic diversity of S.flavescens germplasm resources from different habitats. The results showed that, a total of 311 bands were amplified by 18 pairs of cpSSR primers, and 97.47 alleles were detected, an average of 17.28 bands were amplified per cpSSR primer pair. The average number of alleles (Na) detected was 5.415, effective alleles (Ne) was 4.395, Shannon's information index (I) was 1.535, the diversity index (h) was 0.748, the unbiased diversity index (uh) was 0.832, and the polymorphism information content (PIC) was 0.886, PIC > 0.5 indicated that 18 pairs of cpSSR primers had good polymorphism. The genetic diversity of S.flavescens from different habitats was abundant, with the I was 1.600, the Ne was 4.786, and other genetic diversity indexes of THP germplasms in Wuxiang County of Shanxi Province were all higher, which indicated that THP was the place with rich genetic diversity. Analysis of Molecular Variance (AMOVA) showed that there were great differences among individuals within germplasms, and the genetic variation rate within germplasms was greater than that of inter germplasms. The results of cluster analysis, principal coordinate analysis (PCoA) and genetic structure analysis by STRUCTURE software divided S.flavescens from different habitats into two groups, and the classification results had obvious geographical correlation. The first group of nine S.flavescens germplasms were mainly from Shanxi, Hebei, Shaanxi and Inner Mongolia, and the second group were mainly composed of germplasms from Shandong, Henan, Jiangsu and Anhui. The 18 pairs of cpSSR primers had good applicability for S.flavescens. The genetic diversity of S.flavescens germplasm resources provided a certain theoretical basis for the exploration, utilization and protection and the breeding of excellent varieties of S.flavescens.

Key words: Sophora flavescens Ait., Chloroplast molecular marker, Genetic diversity, Germplasm resources

Table 1

Source information of S.flavescens"

种质Germplasm 来源Source 经度Longitude (E) 纬度Latitude (N) 海拔Altitude (m) 样本量Sample
HD 河北省邯郸市 114°03′~114°40′ 36°20′~36°44′ 1899 10
THP 山西省武乡县土河坪村 112°26′~113°22′ 36°39′~37°08′ 1069 10
XHD 山西省武乡县西河底镇 112°26′~113°22′ 36°39′~37°08′ 1069 10
AG 河北省安国市 115°33′ 38°42′ 32 10
WGS 山西省长治市五谷山村 113°01′~113°40′ 35°50′~37°08′ 976 10
CF 内蒙古赤峰市 116°21′~120°58′ 41°17′~45°24′ 1000 10
TG 山西省太谷区 112°28′~113°01′ 37°03′~37°12′ 808 10
BJ 陕西省宝鸡市 106°18′~108°03′ 33°35′~35°06′ 618 10
YL 陕西省榆林市 109°77′ 38°03′ 1000 10
PY 山东省平邑县 117°37′ 35°30′ 146 10
LY 山东省临沂市 117°24′~119°11′ 34°22′~36°13′ 88 10
LNJZ 辽宁省锦州市 121°51′ 41°31′ 23 10
BZ 安徽省亳州市 115°53′~116°49′ 32°51′~35°05′ 35 10
SY 江苏省沭阳县 118°30′~119°10′ 33°53′~34°25′ 5 10
HNJZ 河南省焦作市 113°26′~113°40′ 35°10′~35°21′ 91 10

Table 2

Information of cpSSR primer"

引物名称
Primer
name		 正向序列
Forward sequence (5′-3′)		 反向序列
Reverse sequence (5′-3′)		 产物长度
Product
length (bp)		 退火温度
Annealing
temperature (℃)
KS1 AAACCGACACGGATTACTCG CCCAATGTACCCTGATACGG 271 54.8
KS2 CCCCCAATCCTTCTTTGATT ATTGGCTGTTCGTCAATTCC 227 52.1
KS3 ATGAGGTTGTGAATCCGAGG CGATTCGATAAACGGCTCAT 246 54.5
KS4 ACAGGATTTGAACCCGTGAC CCTTCCTTATAATTTCATATCCTTCC 223 55.2
KS5 ACCTTCCCGGAGACTGAACT AAAGCTTTTGTTTCGGCTCA 276 58.2
KS6 TCCAATAACCATCCTTCCCTT GAGTTTTCACACCGGAAAGC 213 52.9
KS7 ACTCCTTTGATGGGTGTTGC ACAAAGAAATTCCACGGTCG 228 55.5
KS8 GGAGCACGGAATATCGAAAA CCAGAACCACGATGATTGAA 194 52.3
KS9 AACAGGCTCCGTAAGATCCA CATCTCAGACCTTGCGATTG 268 55.7
KS10 GCCTACGGATCAATCGACAT TGACCACGAAAGTCAAAATGA 229 54.5
KS11 TTTTCACCTCATACGGCTCC TGGTTATATATGGATTGCTAAAATTG 225 54.7
KS12 TTGATGCCTTGATCGAATGA TGAGCCAGGATCGAACTCTC 229 51.1
KS13 GAATCGACAGATCCACCGAC GAATCGACAGATCCACCGAC 250 55.6
KS14 TGAACTAATTGATTTGATTATTTTCCA TTTGGAACTGCCATCCAACT 206 47.9
KS15 TCAAGGGGTTCTCACAAACA ATTCTCGGTTGACAGGGTTG 145 53.6
KS16 AGAAGATTAAGGAACCCCCG GCATGAAACAACTCGAAGCA 245 54.0
KS17 CCGTGGGACTTAAGGAATGA CTCAAAGCAAAGCCAAGGAG 246 54.2
KS18 GGGGGAAACGGATACTCAAT TTCTTCTGCAGTACCTCGCC 270 54.3

Table 3

Size information of cpSSR amplified band"

引物名称
Primer name		 扩增条带数
Number of amplified bands		 扩增条带大小
Amplified band size (bp)
KS1
 22
 244、247、248、249、250、251、253、262、263、264、265、266、267、268、269、270、271、272、273、274、276、278
KS2 11 224、225、226、227、228、229、230、231、236、237、238
KS3 17 240、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257
KS4
 25
 205、206、207、208、209、211、212、213、214、215、216、217、218、219、220、221、222、223、224、231、233、234、235、236、237
KS5
 26
 250、256、257、258、259、260、261、262、263、264、265、267、268、269、270、271、272、273、274、275、276、277、278、279、280、281
KS6 13 207、208、209、210、211、212、213、214、215、216、217、218、219
KS7
 18
 218、219、220、221、222、223、224、225、226、227、228、229、230、231、232、233、234、236
KS8
 24
 197、198、199、200、287、294、295、296、297、298、299、300、301、302、303、305、306、307、308、309、310、311、312、313
KS9 17 256、257、258、259、260、261、262、263、264、265、266、267、268、269、270、271、272
KS10
 25
 202、203、204、205、206、207、208、209、210、211、212、213、214、215、216、217、218、219、220、221、222、223、224、225、226
KS11 15 190、191、192、193、194、195、196、197、198、199、220、221、222、223、224
KS12 12 227、228、229、230、231、232、233、234、235、237、238、239
KS13 13 251、252、253、254、255、256、257、258、259、260、261、262、264
KS14
 18
 197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、215
KS15
 22
 128、129、130、131、132、133、134、135、136、137、138、139、140、141、142、143、144、145、146、169、170、172
KS16 13 238、239、240、241、242、243、244、245、246、247、248、249、250
KS17 9 242、243、244、245、246、247、248、249、250
KS18 11 268、269、270、271、272、273、274、275、276、277、278
总数Total 311
平均Mean 17.28

Table 4

Genetic diversity of cpSSR loci in S.flavescens"

引物名称Primer name Na Ne I h uh PIC
KS1 6.400 5.308 1.739 0.801 0.890 0.915
KS2 4.200 3.405 1.281 0.685 0.761 0.803
KS3 5.867 4.639 1.625 0.773 0.860 0.890
KS4 6.200 5.155 1.690 0.787 0.874 0.917
KS5 6.000 5.093 1.625 0.749 0.833 0.931
KS6 6.267 5.290 1.738 0.807 0.898 0.897
KS7 6.267 5.112 1.712 0.793 0.881 0.888
KS8 5.133 4.040 1.483 0.737 0.819 0.921
KS9 5.067 4.041 1.462 0.731 0.812 0.907
KS10 6.267 5.320 1.730 0.804 0.893 0.940
KS11 4.133 3.203 1.259 0.675 0.750 0.837
KS12 4.667 3.546 1.356 0.698 0.777 0.825
KS13 5.467 4.432 1.559 0.761 0.846 0.858
KS14 5.467 4.281 1.538 0.748 0.831 0.898
KS15 3.667 2.887 1.120 0.620 0.691 0.928
KS16 5.333 4.248 1.532 0.753 0.840 0.862
KS17 5.267 4.181 1.526 0.755 0.840 0.860
KS18 5.800 4.936 1.663 0.793 0.881 0.875
平均Mean 5.415 4.395 1.535 0.748 0.832 0.886

Table 5

Genetic diversity of 15 different germplasms of S.flavescens"

种质Germplasm Na Ne I h uh
HD 5.500 4.542 1.567 0.761 0.874
THP 5.667 4.786 1.600 0.769 0.855
XHD 5.167 4.096 1.470 0.726 0.806
AG 5.000 3.872 1.426 0.714 0.794
WGS 5.167 4.076 1.483 0.737 0.819
CF 5.444 4.387 1.556 0.759 0.844
TG 5.500 4.540 1.566 0.761 0.847
BJ 5.833 4.629 1.591 0.753 0.839
YL 5.556 4.573 1.565 0.754 0.838
PY 5.333 4.385 1.542 0.759 0.843
LY 5.278 4.329 1.502 0.739 0.823
LNJZ 5.556 4.580 1.584 0.768 0.853
BZ 5.278 4.421 1.475 0.713 0.793
SY 5.500 4.405 1.552 0.753 0.837
HNJZ 5.444 4.310 1.555 0.760 0.845
平均Mean 5.415 4.395 1.535 0.748 0.832

Table 6

AMOVA of S.flavescens germplasms"

项目Item 自由度df 平方和SS 均方MS 估计方差Estimated variance 遗传变异率Genetic variation rate (%)
种质间Inter germplasms 14 785.573 56.112 2.611 8
种质内Within germplasms 135 4049.900 29.999 29.999 92
总计Total 149 4835.473 － 32.610 100

Fig.1

Genetic structure analysis of 15 S.flavescens germplasms (a): Estimation number of populations for K ranging (2-10) by ΔK values, (b): K=2, genetic structure of S.flavescens germplasms"

Fig.2

PCoA of 15 S.flavescens germplasms"

Fig.3

UPGMA clustering results of 15 S.flavescens germplasms based on Nei's genetic distance"

[1] 中国科学院《中国植物志》编辑委员会. 中国植物志. 北京: 科学出版社, 1994.
[2] 国家药典委员会. 中国人民共和国药典(一部). 北京: 中国医药科技出版社, 2020:211.
[3] 张晓雯, 李凌宇, 尚海, 等. 苦参碱及其类似物的结构修饰研究进展. 中草药, 2019, 50(23):5892-5900.
[4] 葛淑俊, 孟义江, 李广敏, 等. 我国药用植物遗传多样性研究进展. 中草药, 2006(10):1584-1589.
[5] Li Q, Su X, Ma H, et al. Development of genic SSR marker resources from RNA-seq data in Camellia japonica and their application in the genus Camellia. Scientific Reports, 2021, 11(1):9919.
doi: 10.1038/s41598-021-89350-w
[6] Grover A, Sharma P C. Development and use of molecular markers:past and present. Critical Reviews in Biotechnology, 2016, 36(2):290-302.
doi: 10.3109/07388551.2014.959891 pmid: 25430893
[7] 张征锋, 肖本泽. 基于生物信息学与生物技术开发植物分子标记的研究进展. 分子植物育种, 2009, 7(1):130-136.
[8] 张全芳, 姜明松, 陈峰, 等. 山东省水稻品种(系)的遗传多样性分析. 作物杂志, 2021(4):26-31.
[9] 李琼, 常世豪, 武婷婷, 等. 120份大豆种质资源遗传多样性和亲缘关系分析. 作物杂志, 2021(4):51-58.
[10] Tonti-Filippini J, Nevill P G, Dixon K, et al. What can we do with 1000 plastid genomes?. The Plant Journal, 2017, 90(4):808-818.
doi: 10.1111/tpj.13491 pmid: 28112435
[11] Zhou J, Chen X, Cui Y, et al. Molecular structure and phylogenetic analyses of complete chloroplast genomes of two aristolochia medicinal species. International Journal of Molecular Sciences, 2017, 18(9):1839.
doi: 10.3390/ijms18091839
[12] Yu X Q, Drew B T, Yang J B, et al. Comparative chloroplast genomes of eleven Schima (Theaceae) species:insights into DNA barcoding and phylogeny. PLoS ONE, 2017, 12:e0178026.
doi: 10.1371/journal.pone.0178026
[13] Xu C, Dong W P, Li W Q, et al. Comparative analysis of six Lagerstroemia complete chloroplast genomes. Frontiers in Plant Science, 2017, 8:15.
[14] Gu C H, Tembrock L R, Zheng S Y, et al. The complete chloroplast genome of Catha edulis:a comparative analysis of genome features with related species. International Journal of Molecular Sciences, 2018, 19(2):525.
doi: 10.3390/ijms19020525
[15] 杜久军, 左力辉, 刘易超, 等. 裂叶榆叶绿体基因组及CP-SSR位点分析. 植物遗传资源学报, 2018, 19(6):1187-1196.
[16] 薛宏, 易自力, 肖亮, 等. 芒属植物叶绿体InDel标记的开发与应用. 现代农业科技, 2015(7):153-155,159.
[17] Zhang S A, Gao M Q, Zaitlin D. Molecular linkage mapping and marker-trait associations with NIRPT,a downy mildew resistance gene in Nicotiana langsdorffii. Frontiers in Plant Science, 2012, 3:185.
[18] Breidenbach N, Gailing O, Krutovsky K V. Genetic structure of coast redwood (Sequoia sempervirens [D. Don] Endl.) populations in and outside of the natural distribution range based on nuclear and chloroplast microsatellite markers. PLoS ONE, 2020, 15 (12):e0243556.
doi: 10.1371/journal.pone.0243556
[19] Gichira A W, Avoga S, Li Z Z, et al. Comparative genomics of 11 complete chloroplast genomes of Senecioneae (Asteraceae) species:DNA barcodes and phylogenetics. Botanical Studies, 2019, 60(1):17.
doi: 10.1186/s40529-019-0265-y
[20] 高源, 王大江, 王昆, 等. 新疆野苹果叶绿体DNA变异与遗传进化分析. 植物遗传资源学报, 2020, 21(3):579-587.
[21] Aecyo P, Marques A, Huettel B, et al. Plastome evolution in the Caesalpinia group (Leguminosae) and its application in phylogenomics and populations genetics. Planta, 2021, 254(2):27.
doi: 10.1007/s00425-021-03655-8 pmid: 34236509
[22] 张靖国, 曹玉芬, 陈启亮, 等. 基于叶绿体DNA变异的湖北梨属种质系统进化及遗传多样性分析. 植物遗传资源学报, 2016, 17(4):766-772.
[23] 王荣升, 杨庆文. 基于叶绿体基因多样性的中国水稻起源进化研究. 植物遗传资源学报, 2011, 12(5):686-693.
[24] Shahzadi I, Ahmed R, Hassan A, et al. Optimization of DNA extraction from seeds and fresh leaf tissues of wild marigold (Tagetes minuta) for polymerase chain reaction analysis. Genetics and Molecular Research, 2010, 9(1):386-393.
doi: 10.4238/vol9-1gmr747 pmid: 20309824
[25] 乔永刚, 贺嘉欣, 王勇飞, 等. 药用植物苦参的叶绿体基因组及其特征分析. 药学学报, 2019, 54(11):2106-2112.
[26] Beier S, Thiel T, Münch T, et al. MISA-web:a web server for microsatellite prediction. Bioinformatics, 2017, 33(16):2583-2585.
doi: 10.1093/bioinformatics/btx198
[27] Peakall R, Smouse P E. GenAlEx 6.5:genetic analysis in Excel. Population genetic software for teaching and research-an update. Bioinformatics, 2012, 28(19):2537-2539.
pmid: 22820204
[28] Liu K J, Muse S V. Power Marker:an integrated analysis environment for genetic marker analysis. Bioinformatics, 2005, 21(9):2128-2129.
doi: 10.1093/bioinformatics/bti282
[29] Kumar S, Stecher G, Li M, et al. MEGA X:molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution, 2018, 35(6):1547-1549.
doi: 10.1093/molbev/msy096
[30] Saitou N, Nei M. The neighbor-joining method:a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution, 1987, 4(4):406-425.
doi: 10.1093/oxfordjournals.molbev.a040454 pmid: 3447015
[31] Evanno G, Regnaut S, Goudet J. Detexting the number of clusters of individuals using the software STRUCTURE:a simulation study. Molecular Ecology, 2005, 14(8):2611-2620.
doi: 10.1111/j.1365-294X.2005.02553.x pmid: 15969739
[32] Earl D A, Vonholdt B M. STRUCTURE HARVESTER:a website and program for visualizing STRUCTURE output and implementing the Evanno method. Conservation Genetics Resources, 2012, 4(2):359-361.
doi: 10.1007/s12686-011-9548-7
[33] 王崇, 王连军, 杨新笋, 等. 104个甘薯品种的cpSSR指纹图谱构建及遗传多样性分析. 热带作物学报, 2021, 42(6):1549-1556.
[34] 魏潇, 章秋平, 刘威生, 等. 基于叶绿体SSR单倍型的普通杏演化关系. 植物遗传资源学报, 2018, 19(4):705-712.
[35] Yan Y D, Li X Y, Worth J R P, et al. Development of chloroplast microsatellite markers for Glyptostrobus pensilis (Cupressaceae). Applications in Plant Sciences, 2019, 7(7):e11277.
[36] 李祥栋, 石明, 陆秀娟, 等. 利用叶绿体基因组SSR标记揭示薏苡属种质资源的遗传多样性. 华北农学报, 2019, 34(增1):6-14.
[37] 段永红, 渠云芳, 王长彪, 等. 药用植物苦参SSR-PCR体系的优化与验证. 中国农业大学学报, 2014, 19(5):95-100.
[38] 刘亚令, 耿雅萍, 解潇冬, 等. 基于SSR分子标记的药用黄芪遗传多样性与遗传结构分析. 草地学报, 2019, 27(5):1154-1162.
doi: 10.11733/j.issn.1007-0435.2019.05.006
[39] 程丽莉, 胡广隆, 苏淑钗, 等. 板栗及其近缘种叶绿体SSR遗传多样性分析. 华北农学报, 2015, 30(2):145-149.
doi: 10.7668/hbnxb.2015.02.026
[1] Huang Guibin, Guan Yaobing, Niu Yongqi, Zhou Lilei, Zhao Yongfeng. Comprehensive Evaluation of 12 Major Agronomic Traits of 103 Chickpea Germplasm Resources [J]. Crops, 2023, 39(1): 6-13.
[2] Guo Huanle, Tang Bin, Li Han, Cao Zhongyang, Zeng Qiang, Liu Liangwu, Chen Zhihui. Comprehensive Evaluation of Phenotypic Traits and Classification of Maize Landraces in Hunan Province [J]. Crops, 2022, 38(6): 33-41.
[3] Li Wangsheng, Wang Xueqian, Yin Xilong, Shi Yang, Liu Dali, Tan Wenbo, Xing Wang. Comprehensive Evaluation of Drought Tolerance of Sugar Beet Germplasms at Seedling Stage [J]. Crops, 2022, 38(6): 54-60.
[4] Zhao Xiaoqin, Jia Ruiling, Liu Junxiu, Liu Yanming, Wen Yinhua, Shi Lili, Zhang Juanning, Ma Ning. Agronomic Traits and Genetic Diversity Analysis of 120 Foxtail Millet Germplasms [J]. Crops, 2022, 38(6): 61-69.
[5] Qi Guangxun, Dong Lingchao, Zhang Wei, Yuan Cuiping, Liu Xiaodong, Wang Yingnan, Dong Yingshan, Wang Yumin, Zhao Hongkun. Evaluation of Resistance to Soybean Mosaic Virus Strain 3 (SMV3) in Foreign Soybean Germplasm Resources [J]. Crops, 2022, 38(6): 70-74.
[6] Lei Lei, Guan Zheyun, Cao Shiliang, Wang Yumin, Lin Chunjing, Peng Bao, Liu Peng, Zhao Limei, Li Zhigang, Zhang Chunbao. Classification of Soybean Heterotic Groups Based on SSR Molecular Markers for Yield-Related Traits [J]. Crops, 2022, 38(4): 54-61.
[7] Zhao Yanfei, Wang Jiyong. Germplasm Resource Protection and Breeding Innovation Status and Development Countermeasures——Taking Hunan and Hainan Provinces as Examples [J]. Crops, 2022, 38(2): 1-5.
[8] Li Wenlue, Chen Changli, Luo Xiahong, Liu Tingting, An Xia, Jin Guanrong, Zhu Guanlin. Genetic Diversity Analysis of Phenotypic Characteristics of Kenaf Resources in Zhejiang Province [J]. Crops, 2022, 38(1): 50-55.
[9] Gao Zhanning, Wang Shujie, Feng Hui, Xue Zhenggang, Yang Yongqian, Song Xiaopeng, Jie Yuanfen. Comprehensive Evaluation of Two-Rowed Barley Varieties (Lines) [J]. Crops, 2022, 38(1): 70-76.
[10] Zhang Quanfang, Jiang Mingsong, Chen Feng, Zhu Wenyin, Zhou Xuebiao, Yang Lianqun, Xu Jiandi. Analysis of Genetic Diversity of Rice Varieties (Lines) in Shandong Province [J]. Crops, 2021, 37(4): 26-31.
[11] Li Qiong, Chang Shihao, Wu Tingting, Geng Zhen, Yang Qingchun, Shu Wentao, Li Jinhua, Zhang Donghui, Zhang Baoliang. Analysis of Genetic Diversity and Genetic Relationship for 120 Soybean Germplasms [J]. Crops, 2021, 37(4): 51-58.
[12] Jia Ruiling, Zhao Xiaoqin, Nan Ming, Chen Fu, Liu Yanming, Wei Liping, Liu Junxiu, Ma Ning. Genetic Diversity Analysis and Comprehensive Assessment of Agronomic Traits of 64 Tartary Buckwheat Germplasms [J]. Crops, 2021, 37(3): 19-27.
[13] Xiang Chao, Sun Suli, Zhu Zhendong, Zong Xuxiao, Yang Tao, Liu Rong, Yang Mei, Xian Dongfeng, Yang Xiuyan. Resistance and Molecular Identification to Powdery Mildew of Pea Germplasms in Sichuan [J]. Crops, 2021, 37(3): 51-56.
[14] Pan Xiaoxue, Hu Mingyu, Wang Zhongwei, Wu Hong, Lei Kairong. Evaluation of Agronomic Traits and Cold Tolerance at Germination Stage in Rice (Oryza sativa L.) Germplasms [J]. Crops, 2021, 37(1): 47-53.
[15] Yang Wanjun, Pan Xiangyu, Wang Xiuhua, Wang Lu, Zhao Yan. Genetic Diversity Analysis of Yield and Agronomic Traits of 119 Alfalfa Varieties (Lines) [J]. Crops, 2020, 36(6): 17-22.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!