This study aimed to develop an effective diagnosis for the giant panda rotavirus (GPRV) infection and early recessive infection of giant pandas, and to provide technical support for the quantitative analysis of the virus. Our study was based on the published sequence of the giant panda rotavirus strain vp4 gene in GenBank. A pair of specific primers was designed, and a fast accurate fluorescence quantitative RT-PCR detection method of giant panda rotavirus was established. Primer concentration, annealing temperature and the numbers of cycles then were optimized. In addition, standard curves were constructed and the Fluorescent Quantitative RT-PCR method and the routine RT-PCR were compared. The results show that the newly-developed Fluorescent Quantitative RT-PCR could detect 1.0×100copies/μL of plasmid DNA and its sensitivity was 100 times higher than that of the routine RT-PCR. The specificity and reproducibility test showed that the new method had the advantages of specificity and reproducibility. These results show that the method can be a stable and reliable detection method for GPRV, and could be used for the studies on characteristics of GPRV and for the diagnosingGPRV infection.