一步法快速质粒鉴定方法

One Step Assay for High-throughput Identification

对《分子克隆实验指南》(第三版)中的牙签法小量制备质粒DNA的方法做了改进,减少了 加样次数,免去冰浴、离心等步骤,改进后的方法在琼脂糖凝胶电泳前仅需一步加样和加热过程, 提高了实验效率。同时结合使用多孔道移液器和96孔板,更适合于在高通量筛选中鉴定阳性克 隆。实验对改进后的方法与《分子克隆实验指南》上的方法进行了比较,表明“一步法”的检测效 果、准确率与重复性均有到较好的结果。

The improvement was gained to develop simpler procedure for high-throughput plasmid identification from the original procedure, the reported Toothpick Minipreparation of Plasmid DNA.The sirnplified steps included cutting short the multistep by reagent mixing and cutting out unnecessary extraction steps for identifiction, such as eentrifugation and cooling in ice bath.The results were affirmed by comparing the improved procedure with the reported one and comfirmed for the same good preciseability and repeatabiLity.One Step Assay was performed by only one-step extracting plasmid DNA from bacterial cells before gel electrophoresis. Furthermore effective equipments, such as multichannel pipettes （8 channels ） and 96 well microplates, were applied for high-throughout assay. So this methed is simple,rapid and effective for highthroughpot plasmid identification.