Development of a Real-time RT-PCR Assay for Quantification of Rabbit GAPDH Gene

According to the published GAPDH gene's CDS sequence in GenBank,a pair of primers was designed and synthesized.Through optimization of react system and conditions,the method for detection of GAPDH gene of rabbit by SYBR Green Ⅰreal-time RT-PCR was established successfully.The results show that the lowest copy number for detection of GAPDH gene with this method is 32 copies/μL.and there is a good linear relationship in a wide range from 3.2×101 to 3.2×107 copies/μL(r=0.999).The coefficient of variation(CV) of 5 different concentration of positive plasmids is 1.67% to 4.73% and 2.66% to 8.74% in intra-assay and in inter-assay respectively.The method established in this paper has the advantages of rapidity,high sensibility,high throughput and good repeatability,which provides a methodological basis for quantitative analysis on function gene of rabbit and the expression of some disease gene in rabbit when GAPDH gene is taken as a reference gene.