一种优化的提高质粒产量的提取方法

An optimized method to improve plasmid DNA extraction efficiency

研究旨在优化质粒提取方法,提高质粒产量。对商品化质粒提取试剂盒的提取条件进行了优化,增加了提前小管摇菌和异丙醇-20℃过夜沉淀两个步骤。利用紫外分光光度计、琼脂糖凝胶电泳、双酶切和免疫印迹等方法,对获得质粒的生化特性和生物学功能进行了测定。结果表明,优化的质粒提取方法可提高30%的质粒产量,质粒的生化指标和生物学功能试验结果显示优化方法获得的质粒质量没有下降。

The aim of this study was to increase extraction yield of plasmids DNA by optimizing the extraction method. The protocol of a commercial plasmid extraction kit was optimized here by adding two important steps: pre-incubation of competent cells in small tubes and overnight precipitation of isopropanol at-20 ℃. Then, the biochemical characteristics and biological functions of the extracted plasmids DNA were measured by UV spectrophotometer, agarose gel electrophoresis, double enzyme digestion and western blot. The results showed that the extraction yield of plasmids DNA using the optimized method was increased by 30 percent. And the purity of the plasmids DNA, their status, response to double enzymes digestion and expression efficacy in HEK293 cells and PFF were not negatively affected which indicated that the optimized method was successfully established in this study.