Verification of CcpA gene knockout

To verify whether the CcpA gene was successfully deleted, we designed the primer for PCR. The primer was designed across the upstream and downstream sequences of the CcpA gene, with a size of about 1827bp. Results showed that two (1827bp and 1985bp) PCR products (WT2 and KO2) were produced, then the amplified DNA products of WT2 and KO2 were electrophoresed on a 1% (wt/vol) agarose gel, and sequenced by Shanghai Sangon Biological Engineering Technology and Services Co., Ltd. for further verification. The results of DNA sequencing confirmed that the CcpA deletion strain was successfully constructed.

为验证CcpA基因是否成功敲除，我们设计了PCR扩增引物。该引物跨CcpA基因的上下游序列设计，扩增产物长度约为1827bp。实验结果显示获得两种PCR产物（WT2与KO2），长度分别为1827bp与1985bp；随后将WT2与KO2的扩增DNA产物在1%（wt/vol）琼脂糖凝胶中进行电泳，并交由上海生工生物工程技术服务有限公司（Shanghai Sangon Biological Engineering Technology and Services Co., Ltd.）测序以进一步验证。DNA测序结果证实，CcpA基因敲除菌株构建成功。