桑叶ISSR-PCR反应体系的建立与优化

目的]建立和优化桑叶稳定的ISSR-PCR反应体系和扩增程序。方法]采用5因素4水平的正交试验和单因子梯度优化相结合的方法对最适因素水平进行筛选。结果]适用于桑叶的25μl ISSR-PCR反应体系中各因素的最佳浓度分别为:dNTPs 0.35 mmol/L、Mg2+2.5 mmol/L、Taq酶1.25 U、引物0.4μmol/L、DNA 100 ng。结论]对于桑叶ISSR-PCR试验,一次正交试验基本可以建立满足要求的ISSR-PCR体系。

Establishment and Optimization of ISSR-PCR Reaction System of Morus alba L

Objective] To establish and optimize the stable ISSR-PCR reaction system and amplification program for Morus alba L..Methods] The combination between five factors,four levels orthogonal design and single-factor gradient was used to screen the optimum factors and levels.Result] The suitable PCR reaction system contained 0.35 mmol/L dNTPs,2.5 mmol/L Mg2+,1.25 U Taq DNA polymerase,0.4 μmol/L primer and 100 ng template DNA in total 25 μl reaction solution.Conclusion] For ISSR-PCR test of Morus alba L.,an orthogonal test can basically meet the requirement of ISSR-PCR system.