猪源大肠杆菌floR、CTX-M、mcr-1耐药基因多重PCR检测方法的建立

Development of multiplex PCR for detection of floR, CTX-M , mcr-1 in pig-derived Escherichia coli

关键词:  大肠杆菌  耐药基因  floR  CTX-M  mcr-1  多重PCR

Key words:  Escherichia coli  resistance gene  floR gene  CTX-M gene  mcr-1 gene  multiplex PCR

摘要:

为了建立猪源大肠杆菌对氟苯尼考、β-内酰胺酶类及粘杆菌素耐药基因的多重PCR检测方法；本研究以氟苯尼考耐药基因floR、β-内酰胺耐药基因CTX-M、粘杆菌素耐药基因mcr-1作为目的基因,设计3对特异性引物,通过对多重PCR反应体系及条件的优化,成功建立多重PCR检测方法；该方法的灵敏度为1.46×10⁵CFU/mL,具有高度特异性、敏感性和可重复性；本方法的建立为大肠杆菌中常见耐药基因的快速检测及分子流行病学调查提供了新的手段。

Abstract:

To develop a multiplex PCR for florfenicol, β-Lactamase and colistin resistance genes in pig-derived Escherichia coli; We chose florfenicol resistance gene floR, β-Lactamase resistance gene CTX-M, colistin resistance gene mcr-1 as the target gene in this study;three pairs of specific primers were designed according to the correlative floR,CTX-M,mcr-1 gene sequences from the GenBank. By optimizing the multiplex PCR reaction system and conditions, the multiplex PCR detection method was successfully established. The sensitivity of this method is 1.46×10⁵ CFU/mL, and this method has high specificity, sensitivity and reproducibility; the development of this method provides a new means for rapid detection and molecular epidemiological investigation of common resistance genes in Escherichia coli.