马铃薯SRAP-PCR反应体系优化及验证  

Optimization and Validation of SRAP-PCR Reaction System in Potato

作　　者：张旭[1] 赵兴奎 耿淑娟 彭锁堂[1] 段永红[1] ZHANG Xu;ZHAO Xing-kui;GENG Shu-juan;PENG Suo-tang;DUAN Yong-hong(College of Agriculture,Shanxi Agriculture University,Taigu 030801)

机构地区：[1]山西农业大学农学院,太谷030801

出　　处：《植物遗传资源学报》2018年第5期993-1000,共8页Journal of Plant Genetic Resources

基　　金：山西省自然科学基金(201701D121105);山西省财政支农项目(2015TGSF-10)

摘　　要：为建立马铃薯最佳的SRAP-PCR反应体系,以马铃薯基因组DNA为模板,采用单因素和正交试验相结合的方法,对影响SRAP-PCR反应体系的5个因素(引物浓度、Mg2+浓度、模板DNA用量、d NTPs浓度和Taq DNA聚合酶用量)进行优化,建立马铃薯优化的SRAP-PCR反应体系。结果表明:马铃薯SRAP-PCR最佳反应体系中模板DNA用量为60 ng,Mg2+浓度为1.5 mmol/L,d NTPs浓度为0.25 mmol/L,引物浓度为0.60μmol/L,Taq DNA聚合酶用量为0.75 U。各因素对扩增结果影响依次是:Mg2+浓度>Taq DNA聚合酶用量>模板DNA用量>引物浓度>d NTPs浓度。用6份马铃薯样品DNA对优化体系进行验证,扩增结果清晰稳定,可用于马铃薯遗传多样性分析和遗传图谱构建等研究。The purpose of this study was to optimize the SRAP-PCR reaction system for potato genomic DNAs.By combining the single factor with the orthogonal experimental design,five factors including the concentration of template DNA,Mg^2 ＋,d NTPs,Taq DNA polymerase and primers were tested,in order to establish the optimal SRAPPCR reaction system for potato. The results showed that the optimal PCR reaction mixture for SRAP-PCR contained60 ng template g DNA,1. 5 mmol/L Mg^2 ＋,0. 25 mmol/L d NTPs,0. 60 μmol/L each primer and 0. 75 U Taq DNA polymerase in a 20 μL of reaction volume. The PCR amplification in 6 potato DNA samples showed clear and reproducible polymorphic bands. We observed that the contributions of each factor in PCR reaction were variable（ Mg^2 ＋ Taq DNA polymerase DNA template primer d NTPs）. Thus,the optimized SRAP-PCR system can be applied in analysis of the genetic diversity and construction of the genetic map in potato.

关 键 词：马铃薯 SRAP-PCR 体系优化 单因素试验 正交设计 

分 类 号：S532[农业科学—作物学]