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目的建立优化的华细辛Asarum sieboldiiAFLP反应体系,为华细辛遗传多样性的研究提供技术支持。方法以华细辛叶片为材料,对基因组DNA提取、酶切连接、PCR扩增等操作过程中各关键因素进行分析,筛选最适条件,建立完善的AFLP反应体系。结果建立了优化的华细辛AFLP分析体系:在基因组DNA提取时,提取液中添加巯基乙醇、样品温浴时间为30 min;Tru1Ⅰ和PstⅠ的酶切时间分别为3 h;在扩增反应中,Mg2+终浓度为1.5 mmol/L、TaqDNA聚合酶用量为0.2μL。结论本反应体系可获得良好的AFLP分析结果,可用于华细辛遗传多样性研究。
Objective To establish an optimized Asarum sieboldii AFLP reaction system to provide technical support for the research of genetic diversity of Hua Xia. Methods The key factors of genomic DNA extraction, enzyme digestion, PCR amplification and other key factors were analyzed in the leaves of H. fruticosa. Optimum conditions were selected and a perfect AFLP reaction system was established. Results An optimized Hua Xia AFLP analysis system was established: During the extraction of genomic DNA, mercaptoethanol was added to the extract and the incubation time of the sample was 30 min; the cleavage time of Tru1I and PstI was 3 h; during the amplification reaction, The final concentration of Mg2+ was 1.5 mmol/L, and the amount of Taq DNA polymerase was 0.2 μL. Conclusion This reaction system can obtain good AFLP analysis results and can be used for genetic diversity research of Hua Xia.