Establishment and preliminary application of multiplex PCR for detecting toxin-typing of goat Clostridium perfringens strains / 中国兽医学报

ABSTRACT

According 1o the genome sequences of α.β,e,ι toxins of Clostridium perfringens in GenBank,four pairs of primers targeting α,β,ε,ι toxin genes were designed.After the multiplex PCR reaction condition was optimized,the multiplex PCR for identification and toxintyping of C.perfringens strains was developed.The specificity test showed that the expected fragments of C.perfringens reference strains including A.B,C,D,E five toxin types were amplified successfully from genomic DNA of C.perfringens,respectively.However,a band could not be amplified from Clostidrium novyi and Clostridium septicum as negative control groups.The sensitivity test showed that the limit detection of multiplex PCR was 9.0,17.8,12.2,13.8,18.5 pg DNA of A,B,C,D,E five toxin types C.perfringens,respectively.Repetitive testing showed that the established method had a good repeatability.Nine type A strains of and 1 type C strains of C.Perfringens from 21 clinical samples of dead goat were detected by the multiplex PCR developed in this study.This study establishes the multiple PCR method which not only can detect C.perfringens rapidly but also can identify five toxin types of C.perfringens.