菠萝蜜EST-SSR标记开发及其种质资源遗传多样性分析

Development of EST-SSR Markers in Artocarpus heterophyllus Lam. and Its Genetic Diversity Analysis of Germplasm Resources

[目的]分析菠萝蜜EST-SSR标记的稳定性和多态性揭示能力,并将其用于菠萝蜜种质资源遗传多样性研究.[方法]利用已获得的菠萝蜜cDNA文库中的EST序列,通过搜索SSR序列,设计引物,PCR扩增后,银染检测扩增结果.[结果]设计开发出479对EST-SSR引物,其中399对能在菠萝蜜上扩增出产物,282对具有多态性,多态性扩增引物占58.87％;从中选择60对能够清晰扩增的多态性引物,对64份菠萝蜜种质资源进行遗传多样性分析,60对EST-SSR引物共扩增出188条带,不同引物的扩增条带数在2～11,平均3.13条;其中有168条为多态性条带,多态率为89.36％;每对引物的多态信息含量（PIC）为0.407 7～0.958 9,平均PIC为0.792 2,这说明供试材料具有较高的遗传多样性.系统聚类分析结果表明,在相似系数0.688 1处,可将64份菠萝蜜种质资源分成二天类群,在相似系数0.732 5处,第二大类群又可分为4个亚群.[结论]EST-SSR标记的多态性揭示能力强,用于分析菠萝蜜种质遗传多态性的能力强,其应用前景广阔.

[Objective]To analyze the stability and polymorphism reveal ability of EST-SSR markers in Artocarpus heterophyltus Lam.,and to apply in genetic diversity analysis of germplasm resources.[Method] By using the obtained EST sequence in A.heterophyllus cDNA library,SSR sequences were searched,and primer was designed.After PCR amplification,the amplified results were detected by silver staining method.[Result] A total of 479 pairs of EST-SSR primers were designed.Among them,399 pairs could amplify products in A.heterophyllus DNA,and 282 pairs had polymorphic bands,accounting for 58.87％.Then,60 pairs of polymorphic primers were selected to analyze genetic diversity among 64 germplasms of A.heterophyllus.Sixty pairs of EST-SSR primers produced 188 bands;and the amplified bands per pair of primers ranged from 2 to 11 with an average of 3.13.Among them,there were 168 polymorphic bands,showing a polymorphic rate of 89.36％.Polymorphic information content （PIC） of each pair primers was 0.407 7-0.958 9 with an average PIC of 0.792 2,indicating that the test material had a higher genetic diversity.Cluster analysis showed that 64 A.heterophyllus germplasms were divided into two groups at the similarity coefficient of 0.688 1.The larger group was then divided into four subgroups at the similarity coefficient of 0.732 5.[Conclusion] EST-SSR markers of A.heterophyllus has strong polymorphism reveal ability,and can be used to identify genetic differences among different A.heterophyllus germplasms,showing broad prospects in application.