Analysis of SSR site characteristics of <i>Trifolium repens</i> transcriptome and primer development

ZHANG Tingting; ZHANG Heshan; SONG Kangjie; ZHAO Zeyu; XU Benbo; LIU Yang

doi:10.11829/j.issn.1001-0629.2022-0841

Pratacultural Science > 2023 > 40(9): 2266-2275. > DOI: 10.11829/j.issn.1001-0629.2022-0841

ZHANG T T, ZHANG H S, SONG K J, ZHAO Z Y, XU B B, LIU Y. Analysis of SSR site characteristics of Trifolium repens transcriptome and primer development. Pratacultural Science, 2023, 40(9): 2266-2275. DOI: 10.11829/j.issn.1001-0629.2022-0841

Citation:

PDF (1047 KB)

Analysis of SSR site characteristics of Trifolium repens transcriptome and primer development

1.
College of Life Science, Yangtze University, Jingzhou 434025, Hubei, China
2.
Key Laboratory of Animal Embryo Engineering and Molecular Breeding of Hubei Province / Institute of Animal Husbandry and Veterinary Science, Hubei Academy of Agricultural Sciences, Wuhan 430064, Hubei, China

More Information

Corresponding author:
XU Benbo　E-mail: benboxu@yangtzeu.edu.cn
Received Date: October 26, 2022
Accepted Date: February 02, 2023
Available Online: July 12, 2023

Graphical Abstract

Abstract

Abstract

The main study aim was to comprehensively understand the SSR loci information and sequence characteristics of white clover (Trifolium repens), develop polymorphic SSR primers, and distinguish 20 white clover varieties. An SSR site search was conducted using the sequencing data of the petals transcriptome of different colors of T. repens, and primers were designed in batches. Sixty pairs of primers were randomly selected for polymorphism verification, and primers with high polymorphism were selected for fingerprint construction. A total of 24960 SSRs were obtained, with a frequency of 13.25%, and an average of one SSR locus appeared at approximately 5.29 kb. Six SSR repeating base types were found in the transcriptome of white clover, including the main repeation types: trinucleotides (33.70%), dinucleotides (28.26%), and mononucleotides (25.02%). The other base repeating types were lower, accounting for only 9.01% of the total SSR. The main repeating motifs were A/T (24.85%), AG/CT (16.48%), and AAG/CTT (8.89%) from mononucleotide to trinucleotide, respectively. The number of repetitions of each primitive from mononucleotide to hexanucleotide concentrated between 4 and 21 times, accounting for 98.11% of the total, and the sequence length varied between 12 and 35 bp, accounting for 96.43% of the total sequence length. A total of 18594 primers were yielded from 24960 SSRs, accounting for 74.50%. Among the 60 pairs of random primers, 52 and 17 pairs of could amplify bands and polymorphic primers, accounting for 86.7% and 32.7%, respectively. The fingerprint map of white clover constructed with five pairs of primers with high polymorphism could effectively distinguish the 20 different white clover varieties. The SSR loci of the white clover transcriptome in this study were showed high frequency and rich distribution, with high polymorphism, demonstrating considerable potential for application in the analysis of the genetic diversity, molecular-marker-assisted breeding, and fingerprint profile construction of white clover.
- polymorphic primers,
- fingerprint,
- base repeat characteristics,
- SSR sequence features,
- transcriptome sequencing,
- molecular identification,
- SSR primer design

FullText(HTML)

References (30)

References

[1]	STEINER J J. Molecular phylogenetics of the clover genus (Trifolium-Leguminosae). Molecular Phylogenetics and Evolution, 2006, 39(3): 688-705. doi: 10.1016/j.ympev.2006.01.004
[2]	ALMEIDA J P F, HARTWIG U A, MARCO F, JOSEF N, ANDREAS L. Evidence that P deficiency induces N feedback regulation of symbiotic N₂ fixation in white clover (Trifolium repensL.). Journal of Experimental Botany, 2000(348): 1289-1297.
[3]	AINA R, SGORBATI S, SANTAGOSTINO A, LABRA M, CITTERIO S. Specific hypomethylation of DNA is induced by heavy metals in white clover and industrial hemp. Physiologia Plantarum, 2004, 121(3): 472-480. doi: 10.1111/j.1399-3054.2004.00343.x
[4]	张鹤山, 陈志宏, 田宏, 熊军波, 刘洋. 49份白三叶种质资源遗传多样性的SSR分析. 种子, 2019, 38(11): 1-6. ZHANG H S, CHEN Z H, TIAN H, XIONG J B, LIU Y. SSR analysis on genetic diversity of 49 white clover germplasm resources. Seed, 2019, 38(11): 1-6.
[5]	CHU L, GAO Y P, CHEN L L. Impact of environmental factors on seed germination and seedling emergence of white clover (Trifolium repens L.). Agronomy, 2022, 12(1): 350-359. doi: 10.3390/agronomy12010190
[6]	王子玥, 刘凌云, 刘曼, 常智慧. 基于EST–SSR和种子特征的苇状羊茅遗传分析. 草业科学, 2022, 39(1): 66-74. WANG Z Y, LIU L Y, LIU M, CHANG Z H. Genetic analysis of Festuca arundinacea based on EST-SSR and seed characteristics. Pratacultural Science, 2022, 39(1): 66-74.
[7]	曾令霜, 张晨晨, 张敬, 徐彬. 多年生黑麦草种质SSR分子标记遗传多样性分析. 草业科学, 2022, 39(1): 75-84. ZENG L S, ZHANG C C, ZHANG J, XU B. Genetic diversity analysis of perennial ryegrass germ plasm by SSR molecular markers. Pratacultural Science, 2022, 39(1): 75-84.
[8]	李佳彬, 黄蕾, 张雅楠, 贾媛媛, 田芸芸, 张雷, 党振华. 基于刚毛柽柳转录组测序的EST-SSR标记识别与开发. 草业科学, 2021, 38(4): 630-639. LI J B, HUANG L, ZHANG Y N, JIA Y Y, TIAN Y Y, ZHANG L, DANG Z H. Identification and development of EST-SSR markers based on transcriptome sequencing of Tamarix hispidata. Pratacultural Science, 2021, 38(4): 630-639.
[9]	LI Y C, KOROL A B, FAHIMA T. Microsatellites: Genomic distribution, putative functions and mutational mechanisms: A review. Molecular Ecology, 2002, 11(12): 2453-2465. doi: 10.1046/j.1365-294X.2002.01643.x
[10]	GAITAN-SOLIS E, DUQUE M C, EDWARDS K J, TOHME J. Microsatellite repeats in common bean (Phaseolus vulgaris): Isolation, characterization, and cross-species amplification in Phaseolus ssp. Crop Science, 2002, 42(13): 2128-2136.
[11]	WANG M L, GILLASPIE A G, NEWMAN M L. Transfer of simple sequence repeat (SSR) markers across the legume family for germplasm characterization and evaluation. Plant Genetic Resources, 2004, 2(2): 107-119. doi: 10.1079/PGR200441
[12]	MIAN M A, SAHA M C, HOPKINS A A. Use of tall fescue EST–SSR markers in phylogenetic analysis of cool season forage grasses. Genome Biology, 2005, 48(4): 637-647. doi: 10.1139/g05-029
[13]	MORGANTE M, OLIVIERI A M. PCR-amplified microsatellites as markers in plant genetics. Plant Journal, 1993, 3(4): 175-182.
[14]	JONES E, HUGHES L, DRAYTON M. An SSR and AFLP molecular marker-based genetic map of white clover (Trifolium repens L.). Plant Science, 2003, 165(3): 531-539.
[15]	BARRETT B, GRIFFITHS A, SCHREIBER M, ELLISON C, MERCER C. A microsatellite map of white clover. Theoretical and Applied Genetics, 2004, 109(8): 596-608.
[16]	李莉, 吴永洁, 王元素. 基于SSR标记的贵州野生白三叶遗传多样性分析. 种子, 2017, 36(11): 4-9. LI L, WU Y J, WANG Y S. Genetic diversity analysis of wild white clover in Guizhou based on SSR markers. Seed, 2017, 36(11): 4-9.
[17]	李莉, 吴永洁, 王元素. 基于SSR标记的同一花序贵州野生白三叶遗传多样性分析. 江苏农业科学, 2019, 47(2): 49-53. LI L, WU Y J, WANG Y S. Genetic diversity analysis of Guizhou wild white clover in the same inflorescence based on SSR markers. Jiangsu Agricultural Sciences, 2019, 47(2): 49-53.
[18]	夏岩石, 张润霖, 卢宇鹏, 李荣华, 李光光, 张华, 郭培国. 菜薹品质性状与SSR 标记的关联分析. 园艺学报, 2019, 46(10): 1960-1972. XIA Y S, ZHANG R L, LU Y P, LI R H, LI G G, ZHANG H, GUO P G. Association analysis of SSR markers and quality traits of brassica campestris. Horticultural Plant Journal, 2019, 46(10): 1960-1972.
[19]	KALIA R K, RAI M K, KALIA S, SINGH R, DHAWAN A K. Microsatellite markers: An overview of the recent progress in plants. Euphytica, 2011, 177(3): 309-334. doi: 10.1007/s10681-010-0286-9
[20]	郭聪, 陈燕, 王莹, 谈峰, 柯裴蓓, 沙文锋, 李玉娟. 美国红枫转录组SSR序列分析. 中南林业科技大学学报, 2021, 41(7): 132-141. GUO C, CHEN Y, WANG Y, TAN F, KE P B, SHA W F, LI Y J. SSR sequence analysis of Acer rubrum transcriptome. Journal of Central South University of Forestry & Technology, 2021, 41(7): 132-141.
[21]	倪鹏程. 高加索三叶草EST-SSR引物的开发及异交率的测定. 呼和浩特: 内蒙古农业大学硕士学位论文, 2021. NI P C. Development of EST-SSR primer for caucasian clover and determination of outcrossing rate. Master Theisis. Hohhot: Inner Mongolia Agricultural University, 2021.
[22]	徐舶, 石凤翎, 乌日娜, 乔雨, 王佳. 草原1号杂花苜蓿花蕾转录组SSR序列特征. 西北农林科技大学学报(自然科学版), 2021, 49(8): 17-24. XU B, SHI F L, Wurina, QIAO Y, WANG J. SSR sequence characteristics of transcript from flower bud of grassland No. 1 Hybrid alfalfa. Journal of Northwest A & F University (Natural Science Edtion), 2021, 49(8): 17-24.
[23]	刘艳, 王霞, 贾秀秀, 方强恩. 基于转录组测序的矩镰荚苜蓿SSR和SNP特征分析. 分子植物育种, 1-13. [2023-07-04]. http://kns.cnki.net/kcms/detail/46.1068.s.20220126.0942.002.html. LIU Y, WANG X, JIA X X, FANG Q E. The fingerprints of 13 White Clover (Trifolium repens L.) cultivars based on SSR makers. Molecular Plant Breeding, 1-13. [2023-07-04]. http://kns.cnki.net/kcms/detail/46.1068.s.20220126.0942.002.html.
[24]	刘强, 赵丽, 郭虹霞, 张丽光, 邓妍, 王陆军, 王创云. 玉米野生近缘种大刍草幼苗叶片的转录组SSR分析. 玉米科学, 2020, 28(2): 69-75. LIU Q, ZHAO L, GUO H X, ZHANG L G, DENG Y, WANG L J, WANG C Y. SSR analysis of transcriptome from seedling leaves of wild relatives of maize. Journal of Maize Sciences, 2020, 28(2): 69-75.
[25]	陈亮, 虞富莲, 姚明哲, 吕波, 杨坤, 堵苑苑. 国际植物新品种保护联盟茶树新品种特异性、一致性、稳定性测试指南的制订. 中国农业科学, 2008, 41(8): 2400-2408. CHEN L, YU F L, YAO M Z, LYU B, YANG K, DU Y Y. Preparation of the UPOV guidelines for the conduct of tests for distincness, uniformity and stability-tea plant. Scientia Agricultura Sinica, 2008, 41(8): 2400-2408.
[26]	宋海斌, 崔喜波, 马鸿艳, 朱子成, 栾非时. 基于SSR标记的甜瓜品种(系) DNA指纹图谱库的构建. 中国农业科学, 2012, 45(13): 2676-2689. SONG H B, CUI X B, MA H Y, ZHU Z C, LUAN F S. Construction of DNA fingerprint database based on SSR marker for varieties (lines) of Cucumis melo L. Scientia Agricultura Sinica, 2012, 45(13): 2676-2689.
[27]	管俊娇, 余志慧, 杨晓洪, 王江民, 张鹏, 黄清梅, 张建华. SSR标记在辣椒DUS测试中的应用研究. 植物遗传资源学报, 2019, 20(2): 396-405. GUAN J J, YU Z H, YANG X H, WANG J M, ZHANG P, HUANG Q M, ZHANG J H. Study on the application of SSR markers in pepper (Capsicum annuum L.) DUS testing. Plant Genetic Resources, 2019, 20(2): 396-405.
[28]	张迪, 任立飞, 张璐璐, 张文浩, 王天佐. 基于荧光毛细管电泳技术的苜蓿SSR指纹图谱构建. 中国草地学报, 2020, 42(6): 10-14. ZHANG D, REN L F, ZHANG L L, ZHANG W H, WANG T Z. Construction of SSR fingerprint of alfalfa based on fluorescent capillary electrophoresis. Chinese Journal of Grassland, 2020, 42(6): 10-14.
[29]	蒲小剑, 田久胜, 田新会, 杜文华. 红三叶遗传图谱构建及抗白粉病基因QTL定位. 草业学报, 2018, 27(4): 79-88. PU X J, TIAN J S, TIAN X H, DU W H. construction of the AFLP linkage map and QTL analysis of powdery mildew resistance in red clover. Acta Prataculturae Sinica, 2018, 27(4): 79-88.
[30]	马骢毓, 韩重阳, 马赛男, 李旭旭, 蔡家邦, 汪阳, 张新全, 聂刚. 基于SSR分子标记的13个白三叶(Trifolium repens L.)品种指纹图谱构建. 草地学报, 2021, 29(9): 1892-1899. MA C Y, HAN C Y, MA S N, LI X X, CAI J B, WANG Y, ZHANG X Q, NIE G. The fingerprints of 13 white clover (Trifolium repens L.) cultivars based on SSR markers. Acta Agrestia Sinica, 2021, 29(9): 1892-1899.

Cited By

Get Citation

PDF

XML

Article views (108) PDF downloads (15)

Turn off MathJax

Article Contents

Abstract

References

Analysis of SSR site characteristics of Trifolium repens transcriptome and primer development

Abstract

References

Catalog

Export File

Citation

Format

Content