Objective To analyze the effect of transport and storage conditions on the detection of pathogenic nucleic acid MHV, Reo-3, MNV in laboratory mouse cecal contents samples. Methods MHV, Reo-3 and MNV were mixed with mouse cecal contents and used as reference samples, respectively. They were placed in the lysis buffer of RNA extraction reagent (buffer AVL) or normal saline, and stored at 4℃ and room temperature (22℃-25℃). RNA of these samples was extracted at 1, 2, 3, 7, and 14 days. Then the amount of nucleic acid in samples was detected by real-time fluorescence quantitative PCR. Results A greater decrease of the amount of nucleic acid was observed when the samples were placed in normal saline than that kept in buffer AVL. The amount of nucleic acid in samples stored at 4℃ was found to be higher than that stored at 25℃ room temperature. The amount of nucleic acid in the samples which were kept in buffer AVL at 4℃ for 3 days was higher than 50%, still detectable in the samples kept for 7 days, and undetectable at 14 days. Conclusions Mouse cecal content samples are preferably stored in the lysis buffer of RNA extraction reagent and transported at 4℃ for the detection of MHV, Reo-3, and MNV nucleic acid. It is better to complete the detection test within 3 days.