质粒pBR_(322)电注入E.coliHB_(101)的研究

Study of Plasmid pBR322transformate E. coli HB101 by high-voltage pulses

常用的转化方法是用CaCl_2或CaCl_2/RbCl_2处理受体菌使成感受态。转化率约为10~3—10~6个转化子/μg质粒DNA。近年发展起来的电转化技术可使转化率提高到lC^(?)—10^(10)个转化子/μg质粒DNA。我们以质粒pBR322和大肠杆菌HB101为实验材料,对常用的转化方法和电转化的结果进行了比较,对电转化中获得最佳转化效果和各种参数如电压、脉冲时间、脉冲次数、循环数等进行了探讨。

Former mathods for bacterial transformation are treatment of bacteria with ice-cold solution of CaCl2/RbCl2, which yield 105 -10 6 transformed colonies/μg of supercoiled plasmid very complicated and needing longer time. Method of High-voltages pulses treatment with E. coli and other bacteria has been recently developed to yield a efficiency of 10 9-10 10transformants/μg DNA. Higher transformation efficienties are connected with higher voltages or longer pulses,but there are declines in cell viability. We compared methods for bacterial transformation by using plasmid pBR322 which had stored for a long time and contained about 40% linear plasmid. About 100 times higher transformations were observed by electrotransformation than method by treated with ice-cold Cadi. We also found a appropriate various parameters.