The major histocompatibility complex (MHC) is a large group of genes that encode cell surface glycoproteins and play a crucial role in the acquired immune systems of all teleosts. The study of the MHC is a major focus in the field of fish molecular immunology, and for the auxiliary breeding of disease resistant fish. The great blue-spotted mudskipper () is special for its unique behavioral, physiological, and biochemical characteristics suitable for amphibian life. This study firstly analyzed the sequence characteristics of the gene, then reconstructed the phylogenetic tree and explored the possible molecular evolution. Finally, the mRNA spatial expression pattern of the gene was assessed in healthy tissues, and hepatic and splenic expression patterns were examined after viral mimetic poly(I:C) injection. The results showed that the open reading frame (ORF) was 1,101 bases in length, encoding 366 amino acid residues, with a molecular mass of 41.1 kD and theoretical isoelectric point (pI) of 4.83. The gene included five domains:the leader peptide, α1 domain, α2 domain, α3 domain, and the transmembrane domain. The following secondary protein structure was predicted:-turn for 7.65%, extended strand for 21.31%, and random coils for 33.88%. There were three protein kinase C phosphorylation sites, one casein kinase Ⅱ phosphorylation site, and one N-glycosylation site. Phylogenetic analysis showed that the was most closely related to the . The spatial expression analysis showed that was expressed in all tissues examined, particularly in the kidneys and spleen, followed by the gills and intestines. At 0 h, 6 h, 12 h, 24 h, 36 h, 48 h, and 72 h after stimulation there were no significant changes (>0.05) in expression levels in the control group. During poly(I:C) challenges which were intraperitoneally injected into the liver, mRNA expression level was increased at 6 h (1.63-fold) and the expression increased significantly and was at its maximum at 12 h (<0.01, 5.30-fold). Then, the expression level decreased at 24 h, but was still significantly higher than the control group (<0.05, 2.98-fold). At 36 h, expression level increased again, and was still significantly higher than the control group (<0.01, 4.68-fold). Expression gradually decreased to 2.22-fold and 1.46-fold the control values at 48 h and 72 h, respectively. In spleen tissues, the expression level of mRNA in the experimental group was significantly increased at 6 h (<0.01, 6.25-fold), and the expression level of the experimental group increased significantly to its maximum (<0.01, 9.71-fold) at 12 h. The maximum value expression level was reached earlier than in liver tissue. At 24 h, the expression level also decreased, but was still higher than the control group (<0.05, 3.25-fold). At 36 h, the expression level increased again and was significantly higher than that in the control group (<0.01, 5.81-fold), expression levels gradually returned to normal levels at 48 h and 72 h (1.21-fold and 1.38-fold, respectively). The results suggested that Boleophthalmus pectinirostris against poly(I:C) challenges.