Establishment and Application of Rapid Detection Method for Polymyxin Resistance Gene mcr-1 Based on RPA-LFD Method

Objective: To develop a rapid, efficient and visual method for the detection of bacterial colistin resistance gene mcr-1, so as to provide the basis and convenience for the development of its detection at the grassroots level. Methods: Using recombinase polymerase amplification combined with a lateral flow dipstick (RPA-LFD) assay, supplemented by a hand-held colloidal gold reader. According to the conserved sequence of the mcr-1 gene, a pair of specific RPA primers were designed and synthesized. Through the optimization of the reaction conditions and system, as well as the specificity test, sensitivity test, simulated food sample test and actual sample test, the RPA-LFD assay for visual and quantitative detection of bacterial colistin resistance gene mcr-1 was successfully established. Results: When the primer concentration was 400 nmol/L and the primer ratio was 1:1, the optimal reaction conditions of this method are Mg2+ concentration 14.0 mmol/L, reaction temperature 37 ℃ and reaction time 20 min. The sen