动物造模,荧光定量PCR z-bio 2022-12-28 614

　　Objective To establish a fluorescent quantitative PCR method for mouse cytomegalovirus and make a preliminary application.

　　Methods The conserved sequence of DNA polymerase gene of MCMV Smith strain published by NCBI was selected to design primers and probes, and a fluorescent quantitative PCR method for MCMV was established to verify the specificity, sensitivity, repeatability and stability of the method. MCMV mouse blood samples and 409 mouse blood samples submitted for inspection in 2018.

　　Results The standard curve of the established MCMV real-time PCR method had a Slope of -3.418, an R² value of 0.999, and an amplification efficiency of 96.137%. The lowest quantitatively detectable MCMV content was 47 copies/μL. Using rat cytomegalovirus, simian cytomegalovirus, human herpes simplex virus, pseudorabies virus and frost herpes virus type I as templates, there is no amplification curve, and the specificity is good. Methods The coefficients of variation within and between groups were 0.39%-0.68% and 0.48%-1.01%, respectively, with good repeatability and stability. The maximum dilution of MCMV virus that could be detected in human mouse blood samples was 1:1000 (100.75 TCID50/0.1mL), and 409 mouse blood samples were all negative.

　　Conclusion The established mouse cytomegalovirus fluorescence quantitative PCR method has good sensitivity, specificity and stability, and can effectively detect MCMV in mice. refer to.