动物实验,荧光定量PCR检测 z-bo 2020-08-20 520

　　Objective: To establish a rapid, sensitive and specific TaqMan probe real-time fluorescent quantitative PCR detection method for the quantitative detection of Helicobacter pylori bile.

　　Method: PCR amplification of H. bilis conservative gene P17 sequence, full length ORF435bp, TA clone, construct plasmid standard pMD-HBP17. Through quantitative analysis of the pMD-HBP17 standard, the reaction system was optimized to detect the sensitivity, specificity and repeatability of the TaqMan probe for real-time fluorescent quantitative PCR. 77 clinical samples were tested by the established qPCR method and compared with conventional PCR. Compare test results.

　　Result: The established qPCR detection method showed good linearity and correlation between the plasmid DNA concentration between 108 and 101 copies. The slope of the obtained standard curve is -3.46, the correlation coefficient is 0.999, and the detection sensitivity is 20. The detection rate of 77 clinical samples in the copy was 14.3%, which was higher than the detection rate of normal PCR (7.8%).

　　Conclusion: The established H.bilis qPCR detection method is highly specific, sensitive, and stable, and can be used for quantitative and qualitative detection of Helicobacter pylori.