动物实验,金黄色葡萄球菌,绿脓杆菌,肺炎克雷伯杆菌 z-bo 2020-09-02 433

　　Objective: To establish a multiplex PCR detection method for Staphylococcus aureus, Pseudomonas aeruginosa and Klebsiella pneumoniae, to provide a fast, sensitive and simple method for detecting bacteria in laboratory animals.

　　Method: Design specific primers based on Staphylococcus aureus nuc gene, Pseudomonas LasI gene, Klebsiella pneumoniae PhoE gene, and bacterial universal 16SRNA gene; optimize the concentration and annealing temperature of multiplex PCR primers, establish specificity and specificity Sensitivity detection, multiplex PCR system. The PCR system is used to verify and detect artificially infected specimens and stool samples of laboratory animals, and compare them with traditional detection methods.

　　Result: Multiplex PCR amplified the target bands of Staphylococcus aureus (153bp), Pseudomonas aeruginosa (600bp), Klebsiella pneumoniae (368bp) and universal (520bp). The multiple sensitivity is 10 pg, and the target band of other bacteria is not detected in specific detection. Multiple PCR systems were established to detect different combinations of artificially infected specimens. One case of Pseudomonas aeruginosa was detected in stool 76, which was consistent with the results of conventional detection methods.

　　Conclusion: The multiplex PCR method established in this article has advantages of specificity, sensitivity, simplicity and speed, and provides a reliable method for rapid detection of microorganisms in laboratory animals.