目的:建立缩叶藓属(Ptychomitrium)植物ISSR-PCR反应的最佳体系。方法:利用正交设计实验的方法,采用引物562,10号材料Ptychomitrium gardneri为模板对缩叶藓属植物的ISSR-PCR反应体系中的5种主要因素(Mg2+d、NTPs、引物、模板量、Taq酶量)在4个水平上进行体系优化。结果:确定了缩叶藓属(Ptychomitrium)植物ISSR-PCR反应的最佳体系(25μl)为:Mg2+浓度为3.2mmol/L、dNTPs浓度为0.96mmol/L、引物浓度为1.04μmol/L、模板量10ng、Taq酶量1.5U。利用该体系,采用引物564在16个材料中能有效扩增。结论:这一优化体系的建立为以后缩叶藓属植物的ISSR遗传多样性的分析奠定了基础。
Objective: Develop the most suitable ISSR-PCR system for Ptychomitrum.Method: The orthogonal design was used to optimize inter-simple sequence repeat PCR(ISSR-PCR) amplification system on Ptychomitrum in five factors(Mg2+,dNTPs,primer,DNA template and Taq DNA polymerase) at four levels respectively.Result: The most suitable ISSR-PCR system for Ptychomitrum was established,namely 25μl reaction system containing 3.2mmol/L Mg2+,0.96mmol/L dNTPs,10ng DNA template,1.04μmol/L primer,and 1.5U Taq DNA polymerase.Using the optimized reaction,16 Ptychomitrum materials were easy amplified with primer 564.Conclusion:The result provided a standardizing program for the analysis of interspecies genetic diversity of Ptychomitrum.