三七ISSRPCR反应体系建立及优化

Establishment and Optimization of ISSR-PCR Reaction System for Panax notoginseng

为建立和优化三七ISSR-PCR反应体系,运用正交试验和单因子试验分析模板DNA,Taq DNA聚合酶、引物、Mg2+和dNTPs 5个因子对ISSR-PCR反应影响。结果发现,三七ISSR-PCR 25μL反应体系中5个因子最佳水平:DNA模板为9.6 ng,Taq DNA聚合酶为2.0 u,Primer浓度为1.0 mmol/L,dNTPs浓度为0.68 mmol/L,Mg2+浓度为2.8 mmol/L。研究结果为利用ISSR分子标记技术研究三七提供了理论支持。

Effect for ISSR-PCR reaction of Panax notoginseng on 5 factors （ DNA template, TaqDNA, primer, Mg2 ＋ and dNTPs） were analyzed by orthogonal design and respective factor design for establis- hing and optimizing ISSR-PCR reaction system of P. notoginseng. As a result, the optimal ISSR-PCR reaction system （25 p,L） mixture contained 9.6 ng DNA template, 2. 0 u Taq DNA polymerase, 1.0 mmol/L primer, 2. 8 mmol/L Mg2＋ and 0. 68 mmol/L dNTPs. Theory sustaining for studying P. notoginseng plant by ISSR molecular marker are afforded from this experiment.