利用正交设计优化辣椒SRAP反应体系

Optimization of SRAP-PCR System for Pepper Using Orthogonal Design

为快速建立优化的辣椒SRAP反应体系,利用L9(34)正交表,探讨了Mg2+、dNTPs、引物和模板DNA4种主要反应成分的浓度变化对SRAP扩增结果的影响。结果表明:通过正交设计实验可高效建立优化稳定的辣椒SRAP反应体系;用该法建立的辣椒SRAP优化反应体系为:20μL反应体系中含10×PCRBuffer 2.0μL、Mg2+2.5 mmol/L、dNTPs 0.2 mmol/L,上下游引物各0.4μmol/L、模板DNA10 ng,TaqDNA聚合酶1 U。

To construct the optimum SRAP reaction system of pepper, orthogonal experimental design L9 （3^4 ） was used. And the comprehensive effect of three levels of four main factors（Mg2＋ , dNTPs, primers and DNA template） was assessed. The results showed that by orthogonal design, an optimal and stable SRAP system for pepper was quickly obtained：2. 5 mmol/L Mg2＋ , 0. 2 mmol/L dNTPs, 0. 4 μmol/L primers, 10 ng DNA template and 1 U Taq polymerase within total 20μL reaction solution.