珍稀植物攀枝花苏铁ISSR扩增条件的优化

在利用ISSR技术对攀枝花苏铁（Cycas panzhihuaensis）种质资源遗传多样性进行研究的实验过程中, 对影响PCR扩增效果的一些因素如DNA的提取、模板DNA质量浓度、Taq酶用量、引物用量、dNTP的用量以及退火温度等指标进行筛选和优化。试验结果表明，20μl的反应体系中采用15~25ng 的模板DNA、2.0~2.5mmol/L的Mg2 + 浓度、0.2mmol/L的dNTPs、0.2μmol/L ISSR 引物、0.2U Tag DNA 聚合酶、48 ℃~55 ℃(退火温度随引物不同而定)的复性温度，以及35个循环数为攀枝花苏铁ISSR—PCR 扩增条件的最佳选择。攀枝花苏铁ISSR—PCR 扩增条件的优化为进行攀枝花苏铁种群间遗传分化的研究奠定了基础。

Optimization of ISSR--PCR Amplification in Cycas Panzhihuaensis

The factors influencing ISSR—PCR experiments to analyze the genetic divergence in: cycas parz hihuaensis were investigated. Comparative experiments on the concentrations of template DNA, ISSR primer, Taq DNA polymerase and the annealing temperature were carried out. The results showed that the optimum reaction condition of ISSR experiment was 15~25ng DNA template, 2.0~2.5mmol/Lmg2+, 0.2mmol/LdNTPs, 0.2μmol/L ISSR primer, 0.2U Tag DNA polymerase, annealing temperature of 48~55℃ and 35 cycles in each reaction system of 20μl, which set up a basis for the study of genetic divergence among Cycas panzhihuaensis.