火龙果ISSR优化体系的建立

Optimization of ISSR reaetion system in pitaya

摘要: 为建立稳定的火龙果ISSR-PCR反应体系,采用单因素与正交设计试验相结合的方法,对ISSR反应体系中的主要因素进行了优化筛选。结果表明,20μL ISSR反应体系中各主要成分的最适浓度分别为10×Buffer(含Mg2+)2.0μL,DNA 20.0ng、0.30mmolL-1 dNTP、引物0.35μmolL-1、Taq DNA聚合酶为1.5U,引物(AC)8T的最佳退火温度为54.0℃。利用优化得到的体系对6份火龙果材料进行检验,结果表明优化后的体系适合火龙果的ISSR-PCR反应。

Abstract: In order to establish a stable ISSR-PCR reaction system for pitaya,a single factor experiment and orthogonal method was applied for the optimization.The results showed that the optimum concentrations of different components in 20 μL reaction system were 2.0 μL 10×Buffer(with Mg2+),20.0 ng template DNA,0.30 mmolL-1 dNTP,0.35 μmolL-1 primer and 1.5 U Taq DNA polymerase.The optimized annealing temperature was 54.0℃ for the primer(AC)8T.The established ISSR-PCR system was tested on 6 pitaya samples with satisfactory performance.