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以平菇“唐平26”基因组DNA为模板,以U836为引物,比较了琼脂糖凝胶电泳和不同浓度聚丙烯酰氨凝胶(PAGE)电泳对ISSR-PCR产物的分离效果,并且采用PAGE电泳技术对影响ISSR反应体系的主要因素进行优化。结果表明:以PAGE电泳技术检测平菇ISSR-PCR产物的效果最好,最佳ISSR-PCR反应体系为,在20μL反应体系中,模板20ng,Mg2+浓度为1.5mmol·L~(-1),引物0.8μmol·L~(-1),dNTPs 200μmol·L~(-1),Taq酶1.0U,30个扩增循环。
The isolation of ISSR-PCR products by agarose gel electrophoresis and polyacrylamide gel electrophoresis with different concentrations of U836 as templates was carried out using the genomic DNA of Pleurotus ostreatus “Tang Ping 26” as a template and U836 as a primer PAGE electrophoresis technique was used to optimize the main factors influencing the ISSR reaction system. The results showed that ISSR-PCR was the best method to detect Pleurotus ostreatus by PAGE electrophoresis. The optimal ISSR-PCR reaction system was as follows: 20 ng of template, Mg2 + concentration of 1.5 mmol·L -1, Primer 0.8μmol·L -1, dNTPs 200μmol·L -1, Taq polymerase 1.0U, 30 cycles of amplification.