[Construction of a dual fluorescent reporter system for tracing horizontal transfer of mcr-1-carrying plasmid].

The green fluorescent reporter gene was inserted into the gene interval of polymyxin resistant mcr-1-carrying plasmid (pSH13G841) by homologous recombination of suicide plasmid. At the same time, E. coli J53 with red fluorescent reporter gene was constructed. Using the ability of spontaneous conjugation of drug resistant plasmid (pSH13G841), pSH13G841-GFP plasmid was transferred into J53 RFP bacteria to construct a double fluorescent labeled donor bacterium. The two light-emitting systems could stably and spontaneously express fluorescence without mutual interference. The dual fluorescence report system constructed can be used for visual tracing horizontal transfer of mcr-1-carrying plasmid, the subsequent model can study the colonization, transfer and prognosis of drug-resistant bacteria/drug-resistant genes mcr-1 by using mouse in vivo imaging technology.通过自杀质粒介导的同源重组方法将绿色荧光报告基因（GFP）标记至含mcr-1基因多黏菌素耐药质粒pSH13G841上，形成可发出绿色荧光的质粒pSH13G841-GFP；通过改造大肠J53菌株，构建带有红色荧光报告基因（RFP）标记的大肠杆菌J53-RFP。随后利用pSH13G841-GFP自发接合的能力，将pSH13G841-GFP质粒转化入J53-RFP菌体中，构建出双荧光标记的耐药质粒供体菌模型，能够稳定且自发地发出红色荧光和绿色荧光。构建的双荧光标记系统可用于耐药质粒水平转移的可视化追踪，为动态监测耐药基因mcr-1在生物体内的接合转移规律提供了可能。.