Abstract: A pair of primers was designed according to the sequence of mouse IL-6 and iNOS genes as well as housekeeping gene β-actin,respectively,and recombinant plasmids containing the target gene was constructed as a standard control for IL-6,iNOS and β-actin. Then,the real-time PCR assays based on TaqMan probes for detection of IL-6,iNOS and β-actin genes were established. Every assay possessed a good linear relationship between initial templates and Ct values,and the correlation coefficient of the standard curve was 0.998. It was highly sensitive and had a detection limit of 1×10¹ copies/μL of initial templates. It was highly specific and the fluorescent signals could only be detected by the reaction with cDNA,specific primer and probe for each gene. It was highly reproducible and had a coefficient of variation less than 2 percent for both intra- and inter-assay. The established assays were successfully used to detect IL-6 and iNOS mRNA expression levels in brain and heart tissues from mice experimentally infected with porcine encephalomyocarditis virus (EMCV) GXLC strain. The high sensitivity,specificity and reproducibility of the assays indicated that the TaqMan real-time PCR could be used as an effective tool for detection and quantification of IL-6 and iNOS.

Key words: mouse; IL-6; iNOS; real-time PCR; encephalomyocarditis virus (EMCV)