Abstract: AKT-interacting protien (AKTIP) is a kind of membrane protein, involvimg in the regulation of PI3K/PDK1/Akt pathway. AKTIP gene is still unknown in fish. In the present study, AKTIP gene of Cynoglossus semilaevis was first cloned and full length of cDNA was 1224 bp in size, including 5'-untranslated region (UTR) of 116 bp, 3'-UTR of 117 bp and a complete open reading frame (ORF) of 891 bp, encoding 296 amino acids. Theoretical isoelectric point (PI) of predicted protein was 9.12 and molecular weight was 33.74 kD. Homologous comparison showed that the amino acids sequence of AKTIP in C. semilaevis had a high identity with those of other species. The AKTIP gene was expressed in all tested tissues in the health C. semilaevis and the highest expression was in the ovary. In order to investigate the expression patterns of the AKTIP gene in immune response, the specific expression of AKTIP was performed after Vibrio anguillarum infection and in peripheral blood lymphocytes with different pathogens stimulated. The results showed that the expression of AKTIP gene was up-regulated in the liver, gill, blood, intestinal, head kidney and spleen after V. anguillarum infection. While the AKTIP gene expression in peripheral blood lymphocytes was down-regulated after PGN, LPS, WGP and poly I: C stimulation. The research revealed that AKTIP gene involved in C. semilaevis immune response. These results provide the evidence to explore AKTIP defense mechanism and to control C. semilaevis disease in the further study.