[目的] 筛选最佳的稻瘟菌SSR-PCR反应体系.[方法] 利用正交设计对稻瘟菌SSR-PCR反应体系中的5个因素(Taq DNA聚合酶、Mg2+、模板DNA、dNTP、引物)在4个水平上进行优化试验,筛选出各反应因素的最佳水平并进行退火温度梯度试验筛选最佳的退火温度.[结果] 稻瘟菌SSR-PCR反应的最佳体系为:反应总体积为20 μl,Taq DNA聚合酶1.0 U,Mg2+ 2.0 mmol/L,DNA 100 ng,dNTP 50 μmol/L,引物(ms355~356)0.4 μmol/L,最佳退火温度为58.5 ℃.在此体系下可从稻瘟菌材料基因组DNA中扩增出300 bp左右清晰的目的条带,说明该体系稳定,适用于稻瘟菌基因组DNA的SSR标记.[结论]该研究为稻瘟菌的分子标记、遗传多样性研究和分子育种奠定了基础.%[Objective] The aim was to screen the optimum SSR-PCR reaction system for Magnaporthe oryzae. [Method] The orthogonal design was used to optimize SSR-PCR reaction system for M. oryzae in terms of 5 factors (Taq DNA polymerase, Mg2+, DNA template, dNTP and primer) from 4 levels and the optimum level of each reaction factor were screened. The optimum annealing temperature was screened by annealing temperature gradient experiment. [Result] The optimum SSR-PCR reaction system for M. oryzae was as follows: the total volume was 20 μl, including 1.0 U Taq DNA polymerase, 2.0 mmol/L Mg2+, 100 ng DNA, 50 μmol/L dNTP, 0.4 μmol/L primer (ms355-356), and the optimum annealing temperature was 58.5 ℃. Under the system, a clear target band about 300 bp was amplified from M. oryzae genome DNA, which showed that the system was stable and was suitable for the SSR marker of M. oryzae genome DNA. [Conclusion] The research laid the foundation for the molecular marker and genetic diversity of M. oryzae and molecular breeding.
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