葡萄种质资源ISSR-PCR反应体系的建立与优化

【目的】建立并优化葡萄种质资源稳定的ISSR—PCR体系，为葡萄种质资源研究和种质创新奠定基础。【方法】以新疆22个葡萄种质资源叶片为材料，采用改良CTAB法提取葡萄基因组DNA，并利用单因素法对影响ISSR反应体系中的各个影响因子进行优化。【结果】优化的最佳ISSR—PCR反应体系为：模板DNA30ng，dNTPs0．3mmol／L，引物0．5μmol／L，Taq DNA聚合酶1．0U，反应体系为25．0μL。【结论】优化了新疆葡萄种质资源ISSR-PCR反应体系中的各个影响因子，利用该体系能够扩增获得清晰、稳定的条带，可以用于葡萄种质资源遗传多样性分析。

Optimization of ISSR-PCR reaction system for grape germplasm resources

Objective]The present study was conducted to optimize ISSR-PCR reaction system for grape in order to lay the foundation for research and innovation of Xinjiang grape germplasm resources. Method]Using 22 Xinjiang grape germplasm resources as experimental materials, the modified CTAB method was used to extract grape genomic DNA and the affecting factors in ISSR-PCR reaction system was optimized in single-factor test. Result]The optimizing ISSR-PCR reaction system contained 30 ng of template DNA, 0.3 mmol/L, dNTP, 0.5 μmol/L primer, 1.0 U TaqDNA polymerase in total 25.0 μL of reaction solution. Conclusion]The affecting factors of ISSR-PCR reaction system was optimized and using this system could amplify clear and stable bands, which would be used for analyzing grape genetic diversity and germplasm resources.