拟茎点霉ISSR-PCR反应体系的优化  

Optimization of Inter-simple Sequence Repeat PCR Reaction System for Phomopsis

作　　者：吴鸣谦[1] 闫淑珍[1] 陈双林[1] 

机构地区：[1]南京师范大学生命科学学院,南京210046

出　　处：《生物技术通报》2012年第3期123-127,共5页Biotechnology Bulletin

摘　　要：旨在探讨TaqDNA聚合酶、dNTP、Mg2+、引物和模板DNA等因素对ISSR-PCR扩增的影响,采用单因子试验和正交设计方法相结合建立并优化拟茎点霉反应体系。在此基础上进行引物筛选,同时通过梯度PCR试验,确定引物的最佳退火温度。通过对19个拟茎点霉菌株的检验,结果表明已确立的体系稳定可靠,对不同模板均有较好的适用性和通用性,为利用ISSR技术对拟茎点霉进行遗传分析奠定基础。The single-factor and orthogonal design were used to establish and optimize the ISSR-PCR reaction system for Phomopsis.The effects of 5 factors including Taq DNA polymerase,dNTP,Mg2＋,primers and the template DNA on the PCR reaction were analyzed.On this basis,10 primers were screened with stable amplification and rich polymorphism,the optimal annealing temperature for ISSR-PCR reaction was proposed by gradient PCR.This optimized ISSR reaction system was proved stable and credible for the amplifications for 19 strains of Phomopsis,which would provide the basis for the genetic analysis of Phomopsis.

关 键 词：拟茎点霉 分子标记 正交设计 单因子试验 

分 类 号：Q933[生物学—微生物学]