SNaPshot assay is a single base extension assay which, by extending a base after four primers, displays color of the base through capillary gel electrophoresis to judge the genotype of samples. We aimed to establish a SNaPshot assay to type mouse hepatitis viruses (MHV), including MHV1, MHV3, JHM, and A59. The SNaPshot assay was developed after polymerase chain reaction (PCR) using cDNA from MHV strains two pairs of universal PCR primers and four single-base primers that were designed according to the conserved sequence of the four strains. The PCR products were analyzed by capillary gel electrophoresis. The sensitivity and specificity of the SNaPshot assay were analyzed using gradient dilution method and compared with human Hepatitis B virus (HBV). The accuracy of the SNaPshot assay was analyzed using 41 murine (Mus musculus) serum samples by comparing with ELISA and sequencing the samples containing MHVs. The optimal conditions of SNaPshot assay were that T1﹣T4 primers were 0, 3, 10, and 15 Ts in modification, 4︰6︰5︰10 in concentration, and 16 bp, 19 bp, 26 bp and 31 bp in length. The sensitivity of the SNaPShot assay was 1.25 mg/L (MHV cDNA) (Fig. 3), the specificity was 100% (Fig. 4), and the accuracy was 100% (41/41, Table 3), which was confirmed by ELISA, and the sequencing data. The SNaPshot assay is a sensitive, specific, and accurate method for typing MHV1, MHV3, JHM, and A59.