动物造模 z-bio 2022-10-22 713

　　Objective: To establish a multiplex PCR detection method for Staphylococcus aureus, Pseudomonas aeruginosa and Klebsiella pneumoniae, and to provide a rapid, sensitive and simple method for bacterial detection in experimental animals.

　　Methods: Specific primers were designed according to the nuc gene of Staphylococcus aureus, the LasI gene of Pseudomonas aeruginosa, the PhoE gene of Klebsiella pneumoniae and the bacterial universal 16S rRNA gene; after the optimization of the primer concentration and annealing temperature of multiplex PCR, the specificity and sensitivity were For the detection of sex, a multiplex PCR system was established. The PCR system was used to verify and detect artificially infected specimens and laboratory animal fecal samples, and compared with traditional detection methods.

　　RESULTS: Multiplex PCR amplified the target bands of Staphylococcus aureus (153 bp), Pseudomonas aeruginosa (600 bp), Klebsiella pneumoniae (368 bp) and Universal (520 bp). The multiplex sensitivity was 10 pg, and the specific detection did not detect the band of interest from other bacteria. Different combinations of artificially infected specimens were detected by the established multiplex PCR system, and 1 case of Pseudomonas aeruginosa was detected from 76 feces, which was consistent with the traditional detection method.

　　Conclusion: The multiplex PCR method established in this paper has the advantages of specificity, sensitivity, simplicity and rapidity, and provides a reliable method for the rapid detection of microorganisms in experimental animals.