Objective To study the prevalence of plasmid-mediated quinolone resistance (PMQR) genes in carbapenem-resistant Enterobacteriaceae (CRE) and its resistance mechanism. Methods Clinically isolated CRE strains in a hospital from March 2015 to March 2018 were collected, then identified and performed antimicrobial susceptibility test by VITEK 2 Compact analyzer, carriage of PMQR genes qnrA, qnrB, qnrS, qepA and acc (6')Ib-cr were determined by polymerase chain reaction (PCR) and sequencing, the horizontal transfer of PMQR genes were verified by plasmid conjugation test. Results Resistance rates of carbapenem-resistant Escherichia coli and carbapenem-resistant Klebsiella pneumoniae to quinolones were 100% and 15.56%-33.33% respectively. Detection rate of acc (6')Ib-cr gene was the highest (87.72%), followed by qnrB (77.19%) and qnrS (17.54%), 2 strains (3.51%) carried qnrA gene, qepA gene was not isolated, 84.21% of strains harbored 2 or 3 PMQR genes. PMQR gene was transfected into all the 8 conjugated strains, but minimum inhibitory concentration value of quinolones didn't change significantly. Conclusion The detection rate of PMQR genes in CRE in this hospital is high, but there is a certain sensitivity to quinolones.