梅EST-SSR标记的开发及利用

Development of EST-SSR Markers in Prunus mume and Its Application

利用MISA软件对10 123条梅EST序列进行SSR位点查找,得到含SSR位点的序列935条,SSR位点1 233个,平均每100条EST序列中含有12.18个SSR位点。2核苷酸、3核苷酸重复是最主要的重复类型,分别占35.52%和41.36%。设计了40对EST-SSR引物并进行扩增,有24对引物能扩增出理想的PCR产物,其中17对引物具有较好的扩增多态性。测序后发现13对引物中有73.08%的片段具有相应的SSR位点,对杏DNA指纹中部分谱带的测序结果也证明了是梅扩增出的相应的SSR位点。根据本研究含有SSR位点的测序结果推算,从梅EST中开发真实SSR位点的数目为901。随机选择13对引物对杏和梅进行DNA指纹构建与遗传多样性分析,结果发现,来源于梅的EST-SSR引物在杏中有很高的通用性,这些引物把梅和杏分成了两大群体,说明他们是遗传差异明显的两种植物。

From 10 123 Prunus mume ESTs screened using MISA software,a total of 1 233 SSRs were mined from 935 ESTs with frequency of 12.18 SSR loci distributed over 100 ESTs.Di-and tri nucleotide repeat EST-SSRs were dominant,accounting for 35.52%,and 41.36%,respectively.Forty primer pairs were designed form partial EST-SSRs and used for PCR amplification with 24 primer pairs showing amplifications while 17 were polymorphic.PCR products from 13 EST-SSR primer pairs were selected for sequence analysis where 73.08% of the fragments contain SSRs.EST-SSR motifs were conserved between P.mume and Prunus armeniaca.According to our sequencing results of real SSR loci,there are about 901 SSR loci from these ESTs of P.mume.Thirteen random selection EST-SSR primer pairs were chosen to generate DNA fingerprints and genetic diversity of P.mume and P.armeniaca cultivars.The result showed that the EST-SSR primers designed from P.mume sequences exhibited high transferability between P.mume and P.armeniaca cultivars.Using these EST-SSRs primers,P.mume and P.armeniaca were classified into 2 groups.