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目的:利用生物信息学方法,从虎杖转录组中开发SSRs、SNPs和In Dels标记。方法:从NCBI的SRA数据库下载虎杖RNA-Seq数据,经Trinity拼接后得到unigenes。用MISA软件搜索虎杖unigenes,对得到的SSRs位点信息进行特征分析并利用PRIMER 3软件设计SSRs引物。利用BWA/SAMtools/Var Scan流程开发虎杖SNPs和InDels标记。结果:从虎杖转录组中共搜索得到1 293个SSRs位点,经PRIMER 3设计引物后,共得到948个SSRs标记。同时利用BWA/SAMtools/Var Scan流程获得虎杖SNPs 375 579个,In Dels 13 029个。这些SNPs位点分布在16 186条unigenes上(共14.9 Mb),SNPs位点的密度为1/39.7 bp,核苷酸多样性(θ)为0.02519。6种类型的SNPs变异可以归为置换(233 061,62.1%)和颠换(142 518,37.9%)两类。结论:虎杖基因组积累了丰富的SSRs、SNPs和InDels变异,通过转录组测序获得了大量的虎杖EST-SSRs、SNPs、In Dels标记,为后续虎杖的遗传育种打下了基础。
OBJECTIVE: To develop SSRs, SNPs and In Dels tags from the Huzhu transcriptome using bioinformatics methods. METHODS: RNA-Seq data of Polygonum cuspidatum was downloaded from the NCBI’s SRA database and unigenes were obtained by Trinity splicing. Using MISA software to search Polygonum pubescens unigenes, the characteristic information of the obtained SSRs loci was analyzed and SSRs primers were designed using PRIMER 3 software. Polygonum cuspidatum SNPs and InDels markers were developed using the BWA / SAMtools / Var Scan workflow. Results: A total of 1 293 SSRs loci were obtained from the rhizoma corydalis transcriptome. After the primers were designed by PRIMER 3, 948 SSRs were obtained. At the same time, 375 579 SNPs and 13 029 In Dels were obtained by BWA / SAMtools / Var Scan. These SNPs were located on 16,186 unigenes (14.9 Mb in total), with SNPs at a density of 1 / 39.7 bp and nucleotide diversity (θ) of 0.02519. Six SNP variants can be classified as permutations 233 061,62.1%) and transversion (142 518,37.9%). CONCLUSIONS: The genus Polygonum cuspidatum accumulated a wealth of SSRs, SNPs and InDels variations. A large number of EST-SSRs, SNPs and In Dels markers were obtained from transcriptome sequencing, which laid the foundation for the subsequent genetic breeding of Polygonum cuspidatum.