散斑壳属ISSR-PCR反应条件的优化

Optimization of ISSR-PCR Conditions of Lophodermium

以散斑壳属(Lophodermium)T29、T50、T62、R111菌株为研究材料,对该属ISSR-PCR反应体系中的模板DNA浓度、引物浓度、Mg2+浓度、dNTPs浓度、TaqDNA聚合酶浓度以及退火温度等条件进行优化,寻找出适合此类真菌ISSR-PCR反应的最佳反应体系为15μL反应体系中,模板DNA浓度为4～8ng/μL、引物浓度0.4～0.5μmol/L、Mg2+浓度2.0～2.5mmol/L、dNTPs浓度0.15～0.30mmol/L、TaqDNA聚合酶量1.5～2.5U,退火温度为52℃。

Strain T29 , T50, T62, and R111 of genus of Lophodermium were used as materials in this study to optimized the concentrations of DNA templates, primers, Mg^2＋ , dNTPs, Taq DNA polymerase, the annealing temperature and other conditions in ISSR-PCR system of the genus in order to seek the optimum reaction system of ISSR-PCR suitable for fungi of these kinds. The results showed that the optimal ISSR-PCR system suitable for genus of Lophodermium were as follow： concentration of DNA templates 4- 8 μg/p.L, concentration of primer 0.40-0.50μmol/L, Mg^2＋ 2.0 - 2.5 mmol/L, dTNPS 0.15 - 0.30 mmol/L, Taq DNA polymerase 1.5 - 2.5 U, and the annealing temperature was 52℃.