Based on major capsid protein (MCP) gene sequences of Chinese giant salamander iridovirus (GSIV) inGenBank, specific primers were designed, and the full-length was cloned into the eukaryotic expression vector pcDNA3.1 (+) to construct the recombinant expression vectorpcDNA-MCP. Giant salamander () muscle (GS-M) cells were transfected with the recombinant expressionvector, pcDNA-MCP. At 48 h and 72 h post-transfection, MCP protein expressions in GS-M cells were detectedby indirect immunofluorescence assay. The results showed that the protein expression level at 72 hpost-transfection was significantly higher than that at 48 h post-transfection; western blot assay also confirmed the specificexpression of MCP in GS-M cells at 72 h post-transfection. The eukaryotic plasmid pcDNA-MCP was used as aDNA vaccine to immunize Chinese giant salamanders by injection in dorsal muscle at a dose of 20 g/ind; then, peripheralblood from Chinese giant salamanders in both tested and control groups was collected on day 1, day 3, day 7,day 14, day 21, day 28, and day 35 post-immunization for hemocyte count, classification, and serum-neutralizing antibodytitration. The red and white blood cell counts showed significant increase in numbers of erythrocytes and leukocytesin the peripheral blood of immunized Chinese giant salamanders on day 5 and day 7 post-immunization (±0.76)%,respectively, at day 5 and day 7 post-immunization, and both significantly changed compared with the control group(±︰31.55)]. PCR results revealed thatpcDNA-MCP was distributed in the muscle, liver, spleen, and kidney from day 1 to day 28 post-vaccination. RT-PCRresults revealed that was expressed in all of the above tissues at day 7 and day 28 post-vaccination. A challengetest was conducted at day 28 post-immunization and produced a relative survival of 73.3%. This study provides a fundamentalbasis for the application of the pcDNA-MCP plasmid as a potential DNA vaccine to prevent and control GSIVinfection in Chinese giant salamanders in the future.