索科线虫RAPD反应条件优化

Optimization of reaction condition for RAPD analysis in Mermithidae

以中华卵索线虫为材料，研究索科线虫DNA的提取以及对RAPD的影响因素，包括模板DNA、Mg^2＋、dNTP、引物和Taq聚合酶浓度等，探索最佳反应条件，构建了适于索科线虫RAPD分析的稳定的反应体系．25pL反应体系中，含模板DNA20～40ng，10×PCR缓冲液2．5μL，MgCl2 2．0mmol／L，dNTP150 μmol／L，引物0．3μmol／L，Taq聚合酶1U．实验证明，在对索科线虫进行RAPD分析时，采用优化的反应条件，规范实验操作，使用同厂家同批次的试剂，宴验结果就会具有很好的重复性和稳定性．

This paper studied DNA extraction and factors influencing RADP analysis, ineluding the concentration of DNA template, Mg^2＋ , dNTP, Taq polymerase and primers, in Ovomermis sinensis. The optimized reaction condition of RAPD for Mermithidae in 25 μL reaction volume were as follow： template DNA 20-40 ng, 10 × PCR buffer 2.5 μL, MgCl2 2.0 mmol/L dNTP 15 μmol/L, primer 0. 3 μmol/L, Taq polymerase 1U. As the result, the RAPD can be repeated with optimized reaction condition, the same reagent and standard operation.