LI Rui-jiao
College of Chemistry, Chemical Engineering and Biotechnology,Donghua University,Shanghai 201620,ChinaFENG Jie
Shanghai Laboratory Animal Research Center,Shanghai Quality Monitoring Center of Laboratory Animals,Shanghai 201203XIE Jian-yun
Shanghai Laboratory Animal Research Center,Shanghai Quality Monitoring Center of Laboratory Animals,Shanghai 201203HU Jian-hua
Shanghai Laboratory Animal Research Center,Shanghai Quality Monitoring Center of Laboratory Animals,Shanghai 201203GAO Cheng
Shanghai Laboratory Animal Research Center,Shanghai Quality Monitoring Center of Laboratory Animals,Shanghai 201203ObjectiveTo establish a multiplex PCR procedure for simultaneous detection of H. hepaticus, H. bilis, and H. rodentium, and to preliminarily verify its application value.Methods Three pairs of specific primers for the multiplex PCR procedure were designed according to published H. hepaticus, H. bilis, and H. rodentium 16S rRNA gene sequences, and the reaction conditions of the multiplex PCR were optimized. ResultsThe results showed that all the three pairs of primers could be used to specifically amplify their target bands of 417 bp, 364 bp and 324 bp. The optimal annealing temperature was 52℃, the Mg2+ concentration was 2.0 mM, the dNTP concentration was 200 μΜ, and the concentration of primers was 0.625 μM. The sensitivity of the multiplex PCR for H. hepaticus, H. bilis, H. rodentium was 10 fg. CouclusionsA sensitive, specific, efficient multiplex PCR procedure has been established in this study, providing a good foundation for the detection of H. hepaticus, H. bilis, and H. rodentium.
