高效表达dsRNA大肠杆菌BL21 RNase Ⅲ-的构建

Construction of A Highly Producing DsRNA Escherichia coli Strain BL21 RNase Ⅲ-

为了寻找一种简洁、高效的dsRNA获得方式,降低RNAi技术的生产应用成本。应用PKD46介导的重组系统,将PKD46转入大肠杆菌BL21菌株体内,在L-阿拉伯糖的诱导下,产生重组蛋白,使宿主菌具有同源重组的能力,应用两端具有RNase Ⅲ基因40 bp同源序列的PCR产物对其染色体上的RNase Ⅲ基因进行了基因敲除,并在具有卡那霉素的LB平板上筛选到重组菌株。获得了一株能够利用IPTG诱导高效合成dsRNA的BL21 RNase Ⅲ-菌株,为进一步降低RNAi技术的生产应用成本奠定了基础。

In object to get a simple and efficiently method to RNAi. Using homologous recombination system PKD46, making synthesize dsRNA, cut off the price of using the vector PKD46 transfer into bacteria BL21, and then the recombinant protein was induced by L-arabinose. RNase III was deleted by PCR product with 40 bp homologous sequence in Escherichia coli BL21 with recombinant ability. An RNase III- recombinant bacterium strain that can highly producing dsRNA was obtained. The recombinant bacterium strain would be able to lay foundation for applications of RNAi cheaply.