基于转录组测序的黄枝油杉EST-SSR引物开发  

Development of EST-SSR Primers in Keteleeria calcarea Based on Tran-scriptome Sequencing

作　　者：石远婷 赖柏玟 王晓洁 秦惠珍 柴胜丰[2] Shi Yuanting;Lai Bowen;Wang Xiaojie;Qin Huizhen;Chai Shengfeng(Key Laboratory of Ecology ofRare and Endangered Species and Environmental Protection(Ministry ofEducation),Guangxi Normal University,Guilin,541006;Guangxi Institute of Botany,Guangxi Zhuang Autonomous Region and Chinese Academy of Sciences,Guilin,541006)

机构地区：[1]广西师范大学,珍稀濒危动植物生态与环境保护教育部重点实验室,桂林541006 [2]广西壮族自治区中国科学院广西植物研究所,桂林541006

出　　处：《分子植物育种》2024年第7期2257-2264,共8页Molecular Plant Breeding

基　　金：广西自然科学基金项目(2018GXNSFAA281015)资助。

摘　　要：黄枝油杉是中国特有的珍稀濒危树种,具有较高的生态、经济和科研价值。本研究基于转录组测序结果对黄枝油杉进行EST-SSR标记开发,使用MISA软件对黄枝油杉132 191条unigenes进行SSR搜索,共获得6 903个SSR位点,出现频率为5.22%,平均12.015 kb出现一个SSR位点。SSR重复类型以单核苷酸重复为主,二核苷酸重复和三核苷酸重复次之,所占比例分别55.15%、17.14%、17.07%。采用SSR分子标记筛选出225对扩增清晰且符合预期片段大小的引物,有效扩增率为69.23%,经复筛最终获得9对具有多态性SSR引物。对1个野生黄枝油杉种群25个样品进行多样性检测,所有位点均具有多态性。在9对多态性SSR位点中,等位基因数(Na)、有效等位基因数(Ne)、Shannon's指数(I)、观测杂合度(Ho)、期望杂合度(He)、多态信息含量(PIC)分别为5.111、2.668、0.963、0.456、0.461、0.428。结果表明,9对SSR引物遗传多样性处于中等水平,基于黄枝油杉转录组序列的SSR标记开发是可行有效的,为开展黄枝油杉遗传多样性分析及辅助育种等研究提供参考。Keteleeria calcarea Cheng et L.K.Fu,which is an endemic and endangered species to China,has high ecological,economic and scientific research value.The aim of this study is to develop EST-SSR molecular mark-ers of Keteleeria calcarea based on transcriptome sequencing date.Searching for the SSR loci from 132191 uni-genes of K.calcarea with MISA software,a total 6903 SSRs were obtained,accounting for 5.22%of the total uni-genes,averagely one SSR per 12.015 kb.The repeat type of the SSR loci in K.calcarea was dominated by mononucleotide repeats,followed by dinucleotide repeats and trinucleotide repeats,accounting for 55.15%,17.14%and 17.07%,respectively.By SSR molecular markers,225 pairs of primers were detected,which accounted for 69.23%of the total primers.These 225 pairs of primers amplified a clear fragment with expected size,9 pairs of which with polymorphism were identified after re-screening.These markers were tested on 25 individuals from a wild K.calcarea population,which showed that all of these loci were polymorphic.Among the 9 pairs of polymorphic SSR locus,the average allele number(Na),effective alleles(Ne),Shannon's index(I),observed heterozygosi-ty(Ho),expected heterozygosity(He),and polymorphic information content(PIC)were 5.111,2.668,0.963,0.456,0.461 and 0.428,respectively.The results indicate that the genetic diversity of 9 pairs of EST-SSRs primers are at a moderate level,and the development of EST-SSR markers based on transcriptome sequences are feasible and ef-fective,which will contribute to providing an experimental basis for genetic diversity analysis and facilitating the breeding programs of K.calcarea in the future.

关 键 词：黄枝油杉 转录组 遗传多样性 SSR 引物开发 

分 类 号：S791.15[农业科学—林木遗传育种]