铜鱼ISSR-PCR反应体系的正交设计优化

采用正交设计方法,对影响铜鱼(Coreius heterodon(Bleeker))ISSR-PCR扩增结果的5个因素,如Mg2+、dNTP、引物、TaqDNA聚合酶和模板DNA的浓度,在4个水平上进行了比较优化,建立了适合铜鱼的最佳ISSR-PCR反应体系:在25μL反应体系中含有1.5 mmol/L Mg2+,0.16 mmol/L dNTP,0.2μmol/L引物,1UTaqDNA聚合酶和140 ng模板DNA.检验结果表明,采用该反应体系对铜鱼进行ISSR-PCR扩增可获得多态性高,稳定性和重复性好的电泳条带.     

铜鱼ISSR-PCR反应体系的正交设计优化

采用正交设计方法,对影响铜鱼(Coreius heterodon(Bleeker))ISSR-PCR扩增结果的5个因素,如Mg~(2+)、dNTP、引物、Taq DNA聚合酶和模板DNA的浓度,在4个水平上进行了比较优化,建立了适合铜鱼的最佳ISSR-PCR反应体系:在25μL反应体系中含有1.5 mmol/L Mg~(2+),0.16 mmol/L dNTP,0.2 μmol/L引物,lU Taq DNA聚合酶和140 ng模板DNA.检验结果表明,采用该反应体系对铜鱼进行ISSR-PCR扩增可获得多态性高,稳定性和重复性好的电泳条带.

Abstract：

The method of orthogonal design was used to optimize five factors,i.e.concentration of Mg~(2+),dNTP,primer,Taq DNA polymerase and DNA temple,which affect ISSR-PCR amplification results of Coreius heterodon(Bleeker),each at four levels,and an optimal ISSR-PCR reaction system for C.heter-odon(Bleeker)was established:an ISSR-PCR reaction system with a total volume of 25 μL containing 1.5mmol/L Mg~(2+),0.16 mmol/L dNTP,0.2μmol/L primer,1U Taq DNA polymerase and 140 ng DNA tem-pie.The testing results of the reaction system confirmed that it can produce stable and reproducible electrophoretic bands with high polymorphism of ISSR-PCR for C.heterodon(Bleeker).