齿裂菌属和皮下盘菌属ISSR-PCR最佳反应条件的研究  

Optimization of ISSR-PCR Reaction Conditions for Coccomyces and Hypoderma

作　　者：于红梅[1] 林英任[1] 韩加军[2] 刘艳兵[2] 唐燕平[1] 

机构地区：[1]安徽农业大学林学与园林学院,合肥230036 [2]安徽农业大学生命科学学院,合肥230036

出　　处：《菌物研究》2007年第4期225-228,232,共5页Journal of Fungal Research

基　　金：国家自然科学基金项目(30370011;30470014);安徽省自然科学基金项目(070411005)

摘　　要：在利用ISSR技术分析齿裂菌属和皮下盘菌属遗传多样性的研究中,为获得条带清晰、重复性好的ISSR扩增结果,对影响ISSR-PCR的条件进行了筛选,确定了此类菌物ISSR-PCR反应的最适宜条件:在15μLPCR反应体系中,10倍Taq酶缓冲液1.5μL,DNA模板8ng/μL,MgCl22.5mmol/L,dNTP0.15mmol/L,引物浓度0.4μmol/L,Taq酶1.00U,ddH2O9.0μL。最佳退火温度因不同的引物而定,最佳循环次数为35次。The factors which affect the ISSR analysis in the study of the genetic diversity of Coccomyces De Not. and Hypoderma De Not., such as concentrations of template DNA, Taq DNA polymerase, Mg^2 ＋, primer and dNTP, annealing temperature, cycling times, etc. were filtrated. The optimal ISSR - PCR conditions in the experiments were as the following： 10 × Taq buffer 1.5μL, template DNA 8 ng/μL, MgCl2 2.5 mmol/L, dNTP 0.15 mmol/L, primer concentration 0.4 mmol/L, Taq DNA pelymerase 1.00 U, and ddH20 9.0 μL in total 15 μL reaction volumes. Optimal annealing temperature is decided by different primers. Optimal circular time is thirty-five. The amplification system can give clear and reproducible banding patterns.

关 键 词：齿裂菌属 皮下盘菌属 ISSR—PER反应条件 优化 

分 类 号：Q75[生物学—分子生物学] Q949.325