随机扩增多态性DNA最佳模板提取方法的探讨

Study of Optimal Method to Extract Template for Random Amplified Polymorphic DNA

目的 获得最佳模板提取方法用于随机扩增多态性DNA(RAPD)。方法 对 4种方法 (挑丝法、沉淀法、水清法和裂清法 )提取的模板DNA进行琼脂糖电泳、紫外线扫描及随机扩增多态性DNA ,比较其片段大小、纯度及RAPD指纹图差异。结果 挑丝法和沉淀法均可提取出大于 2 3kb且纯度较高的模板DNA ,二者指纹图完全一致且均有 2～ 4kb的较大片段。水清法和裂清法提取的模板有弥散、碎裂的DNA并纯度较差 ,指纹图上仅有 6 0 0～ 2 0 0 0bp的较小片段。 结论 用挑丝法或沉淀法提取的模板DNA最适于进行RAPD。

Objective To obtain optimum template for random amplified polymorphic DNA (RAPD). Method 4 different methods (rod winding, centrifuge, water-supernatant and lysis-supernatant) to extract template DNA were used, the length and purity of template DNA and its RAPD profile were observed by agarose slab gel electrophorse and spectrophotometric scan assays and RAPD reaction. Result The template DNA (>23 kb) with high purity and the same RAPD profile with 2～4 kb DNA fragments were obtained by both rod winding and centrifuge method. However, the template DNA (4 kb and 2 kb respectively) with broken and semar and low purity was extracted by method of water supernatant and lysis-supernatant, and 600～2 000 bp DNA fragments were seen in the similar RAPD profile. Conclusion Template DNA extracted by rod winding or centrifuge method was the optimization for RAPD.