SRAP体系中苔藓植物遗传多样性分析中的正交优化及初步应用

Optimization of orthogonal design and initial application in genetic diversity analysis for bryophytes in SRAP system

以黄灰藓为材料,通过正交法优化SRAP-PCR反应条件,并在此基础上对11种苔藓植物进行遗传多样性分析。结果表明:苔藓植物SRAP反应(25μL体系)的最佳条件为:40 ng DNA模板,2.5 mmol/LMg^(2+),0.3 mmol/L dNTP,15 pmol引物,1.5 U Taq酶。用30对引物组合进行筛选,有5对引物组合扩增条带清晰,重复性好,共扩增出65条带,其中多态性条带63条,多态性比率为96.9%。通过SPSS11.5分析软件对扩增结果进行聚类分析,结果与形态学分类基本一致,说明SRAP技术可用于苔藓植物的遗传多样性研究。

An optimal protocol of SRAP-PCR was established by orthogonal method for Hypnum pallescens. Based on it, genetic diversity of 11 bryophyte species was analyzed in the present study. The results showed that the optimal protocol was accomplished in 25μL reaction volumes containing 40ng template DNA, 2.5 mmol/L MgCI2,0.3 mmol/ L dNTPs,15 pmol/L SRAP primer and 1.5 unit Tag DNA polymerase. Five pairs of primers were selected from thirty ones to generate sixty-five DNA bands which were clear and repetitive, sixty-three of which were polymorphic. Polymorphic percentage was 96.9%. A dendrogram was established based on Hierarchical cluster analysis by SPSS 11.5, which was almost the same as morphological studies,showing that the SRAP technology could be used to analyze the genetic diversity of bryophytes.