利用正交设计建立与优化北美驼绒藜ISSR-PCR反应体系

利用正交试验设计的方法，对北美驼绒藜ISSR-PCR反应的5因素(Taq酶、dNTP、引物、Mg²⁺和模板DNA)4水平进行试验，试验结果运用MINITAB软件进行分析，建立了适合北美驼绒藜的既稳定又谱带多的ISSR-PCR最佳反应体系，即20 μL的反应体系中含有1×buffer，1.5 U Taq酶，0.2 mmol·L^-1 dNTP，0.5 μmol·L^-1引物，2.5 mmol·L^-1 Mg²⁺和10 ng模板DNA。这一优化的ISSR-PCR反应体系的建立，为今后利用ISSR技术进行驼绒藜属植物种质资源分类、遗传图谱构建和遗传变异奠定了技术基础。

Establishment and Optimization of ISSR Reaction System for Winterfat Using Orthogonal Design

The orthogonal design was used to optimize ISSR amplification system of winterfat in five factors (Taq DNA polymerase, dNTP, primer, Mg²⁺ and DNA templet) at four levels respectively. The data were analyzed by software MINITAB. As a result, a satisfactory ISSR reaction system for winterfat with desirable repeatability and polymorphic bands was established. In a total volume of 20 μL ISSR-PCR system, it contained 1×buffer, 1.5 U Taq DNA polymerase, 0.2 mmol·L^-1 dNTP, 0.5 μmol·L^-1 primer, 2.5 mmol·L^-1 Mg²⁺ and 10 ng template DNA. This optimized ISSR reaction system would provide the basis for the analysis of diversity, map construction and genetic variation of important traits of Ceratoides with ISSR markers.