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[目的]建立适用于蒙药阿给(Artemisia frigidaWilld.)的ISSR反应体系,为ISSR标记技术在蒙药阿给遗传多样性研究中的应用奠定技术基础。[方法]利用正交试验设计,从Mg2+、dNTP、TaqDNA聚合酶和引物浓度4因素3水平,对蒙药阿给ISSR-PCR反应体系进行优化分析,并在此基础上对PCR的循环次数及退火温度进行试验。[结果]在20μl反应体系中,Mg2+为1.5mmol/L,dNTP为0.25mmol/L,TaqDNA聚合酶为1.5U,引物为0.75μmol/L时,且ISSR最适的循环次数为40,最适退火温度为56.0℃,扩增效果最好。[结论]筛选出的反应体系适用于蒙药阿给的ISSR扩增。
[Objective] The research aimed to establish ISSR reaction system suitable for Artemisia frigidaWilld., Which laid the technical foundation for the application of ISSR marker in the genetic diversity of Mongolian medicine. [Method] The orthogonal design was used to optimize the ISSR-PCR reaction system of Mongolian medicine Yaman from Mg2 +, dNTP, Taq DNA polymerase and primer concentration of 4 factors and 3, and on this basis, the number of cycles of PCR and annealing Temperature test. [Result] In the 20μl reaction system, the optimal number of cycles of ISSR was 1.5mmol / L, dNTP was 0.25mmol / L, Taq DNA polymerase was 1.5U, primer was 0.75μmol / L, Annealing temperature of 56.0 ℃, amplification best. [Conclusion] The screened reaction system was suitable for ISSR amplification of Mongolia Mongolian medicine.