CLONING AND EXPRESSION ANALYSIS OF Pt-ditt4l IN SWIMMING CRAB (PORTUNUS TRITUBERCULATUS)

The DNA Damage Inducible Transcript like 4 cDNA of swimming crab Portunus trituberculatus was cloned by RACE method to clarify the structure and function of Pt-ditt4l gene. The gene sequence of Pt-ddit4l was 2727 bp long, containing an ORF of 507 bp to encode 168 amino acids. The molecular mass of Pt-ddit4l was 19.14 kD, and has a highly conserved RTP801_C domain at the C-terminal. Blast similarly alignment and phylogentic tree analysis showed that Pt-ddit4l gene had a 60% similarity with Litopenaeus vannamei ddit4l gene, and first clustered with Litopenaeus vannamei. The expression of Pt-ddit4l at different developmental stages of swimming crab showed that it played a role in early development. The Pt-ddit4l was expressed in all tested tissues with the highest expression in hepatopancreas and the lowest in hemocytes. The Vibrio parahaemolyticus infection experiment showed that the expression of Pt-ddit4l in hemocytes increased gradually and then decreased, reaching a peak at 24h, which was 8.85-fold that of the control group (P<0.05). However, the trend was down-regulated with a minimum at 72h in hepatopancreas, which was only 1/40 of the control group (P<0.05). After RNA interference, the expression level of Pt-ddit4l was down-regulated within 6−24h, and the mortality rate of the infected swimming crab with dsddit4l injection was much higher than that of the control group. This study provides a reference for further understanding the physiological function of ddit4l gene in P. trituberculatus.