Establishment of real-time PCR method for detecting HSV-2
To develop a real-time PCR method for herpes simplex virus-2 (HSV-2). Methods The recombinant plasmid pMD18-T-gG was constructed as HSV-2 DNA standard for quantitative analysis. According to the principle of TaqMan MGB technique, the primers and probe targeted at HSV-2-gG gene fraction were designed and applied to detect HSV-2. Then the PCR reaction system was optimized and evaluated. Results The recombinant plasmid was constructed successfully. The real-time PCR method for HSV-2 using TaqMan MGB technique was established and the assay showed good reproducibility (intra-assay coefficients of variation was 2.29% and inter-assay coefficients of variation was 4.76% ) , good specificity (it can distinguish HSV-2 from other pathogenic microorganism) ,good linear range (4.75×10~1-4.75×10~6 copies/μl, r = 0.998) and sensitivity reached 47.5 copies/μl. At the same time, this new established method had more advantages than the ELISA method. Conclusion A novel real-time PCR detecting method for HSV-2 has been developed, which can be used in clinical detection.