果梅ISSR-PCR反应体系的建立和优化

Establishment and Optimization of an ISSR-PCR Reaction System in Japanese Apricot (Prunus mume Sieb.et Zucc.)

以果梅品种桃梅为试材，通过单因子试验分别研究了DNA模板浓度、引物浓度、Mg^2＋浓度、dNTPs浓度、退火温度及循环数等对果梅ISSR-PCR反应的影响．建立了果梅ISSR-PCR的最佳反应体系：20μL反应体系中含10×buffer2μL，2．5mmol／LMgCl2，0．2mmol／LdNTPs，0．32μmol／L引物，20～80ng模板DNA，1UTaq DNA聚合酶．利用优化的反应体系，对19个果梅品种进行体系稳定性检测，结果表明该反应体系的重复性和稳定性良好．

In order to use ISSR markers in studies of genetic diversity, cultivar identification and assistant breeding of Japanese apricot （Prunus mume Sieb. et Zucc. ）, the factors affecting ISSR results of Japanese apricot were studied, such as concentrations of template DNA, Mg^2＋ , dNTPs, primer and Taq polymerase and annealing temperature. An optimal reaction system （20μL） was established, i. e. 2.0μL 10× buffer, 2.5 mmol/L MgCl2, 0.2 mmol/L dNTPs, 0.32 μmol/L primer, 20-80 ng template DNA and 1 unit of DNA polymerase. Nineteen cultivars of Japanese apricot were used to test the stabilization of the optimized reaction system. The results indicated that the optimized reaction system of ISSR-PCR of Japanese apricot was stable and repeatable.