论文部分内容阅读
目的为了获得药用植物黄芪SSR-PCR反应的最优体系。方法以山西浑源黄芪为试验材料,采用单因素实验的方法,对影响SSR扩增结果的SSR-PCR反应体系中的5种反应组分(模板DNA、dNTP、TaqDNA聚合酶、引物及退火温度)进行了探索。结果在20μl反应体系中,模板DNA的用量为60 ng,引物的最适浓度为0.6μmol/L,dNTP浓度为0.1 mmol/L、TaqDNA聚合酶浓度为0.05 U/μl、退火温度为57℃为最适反应体系。利用此反应体系对不同来源(山西、黑龙江、辽宁、河北、甘肃)的黄芪进行PCR扩增及琼脂糖凝胶电泳检测,扩增结果清晰且谱带多态性丰富。结论成功建立了药用植物黄芪的SSR-PCR最优体系,并利用筛选出的引物检测出不同产地黄芪具有很高的多态性,为今后黄芪种质资源的分子评价奠定了技术基础。
Objective To obtain the optimal system for the SSR-PCR reaction of medicinal plant Astragalus. Methods In this study, we selected the four kinds of reaction components (template DNA, dNTP, TaqDNA polymerase, primer and annealing temperature) in the SSR-PCR reaction system that affected the SSR amplification results. ) Has been explored. Results In the 20μl reaction system, the amount of template DNA was 60 ng, the optimum concentration of primer was 0.6μmol / L, the concentration of dNTP was 0.1 mmol / L, the concentration of Taq DNA polymerase was 0.05 U / μl, the annealing temperature was 57 ℃ The most appropriate reaction system. Astragalus from different sources (Shanxi, Heilongjiang, Liaoning, Hebei, Gansu) were amplified by PCR and detected by agarose gel electrophoresis using this reaction system. The amplified results were clear and the bands were rich in polymorphic bands. Conclusion The optimum system of SSR-PCR for medicinal plant Astragalus was successfully established. Using the selected primers, the high polymorphism of Astragalus membranaceus in different areas was detected, which laid the technical foundation for the future molecular evaluation of Astragalus germplasm resources.