慈姑种质资源ISSR遗传多样性分析

Genetic diversity of arrowhead (Sagittaria trifolia) resources based on ISSR

利用ISSR分子标记技术对慈姑种质资源进行分析,以揭示不同来源慈姑间的遗传多样性和亲缘关系。采用正交试验对慈姑ISSR-PCR反应体系中各个影响因子进行优化,利用优化后的体系对ISSR通用引物进行筛选,选择出条带清晰、多态性高的引物,并以41个不同地理来源的慈姑为材料进行扩增。优化后的反应体系为DNA 30 ng、Mg2.5 mmol·L^(-1)、dNTPs 0.8 mmol·L^(-1)、引物1μmol·L^(-1)和Taq DNA聚合酶1.5 U。基于优化体系从100条ISSR引物中筛选出16条用于慈姑分析。对慈姑材料进行ISSR-PCR扩增,共获得条带155条,其中121条多态性条带,多态性比率为78.1%,观测等位基因数(N)为1.780 6,有效等位基因数(N)为1.410 3, Nei’s基因多样性指数(H)为0.243 6, Shannon’s信息指数(I)为0.369 9。不同慈姑的遗传相似性系数在0.796~0.923之间,当相似性系数为0.820时,聚类分析可将慈姑材料聚成5个类群。

The ISSR molecular marker technique was used to analyze the genetic diversity and genetic relationship of arrowhead(Sagittaria trifolia) germplasms from different regions. The orthogonal experiment was used to optimize the various influencing factors in the ISSR-PCR reaction system. Using this system, primers which had clear bands and high polymorphism were selected. Then 41 varieties of different geographical origins were analyzed. The optimal ISSR-PCR reaction system was: DNA 30 ng, Mg2.5 mmol·L^(-1), dNTPs 0.8 mmol·L^(-1), primer 1 μmol·L^(-1)and Taq DNA polymerase1.5 U. 16 primers suitable for arrowhead ISSR analysis were screened from 100 primers.A total of 155 bands were amplified, of which 121 were polymorphic, and the polymorphic ratio was 78.1%. The observed number of alleles(N) was 1.780 6, the effective number of alleles(N) was 1.410 3, Nei’s gene diversity index(H) was 0.243 6 and Shannon’s information index(I) was 0.369 9. The genetic similarity coefficient was 0.796-0.923. When the similarity coefficient was 0.820, the arrowhead materials could be clustered into five groups by clustering analysis.