| Citation: |
PAN Liling, HUANG Guiju, CHENG Shuying, WANG Xiaoning, YU Dahui. cDNA cloning, characterization and challenge-based expression profiles of cathepsin D in winged pearl oyster Pteria penguin[J]. South China Fisheries Science, 2012, 8(2): 22-29. DOI: 10.3969/j.issn.2095-0780.2012.02.004
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We cloned the cathepsin D gene of winged pearl oyster(Pteria penguin)and conducted preliminary studies on its expression profiles. The cDNA sequence of cathepisn D was obtained from P.penguin (named as pgCTSD) by using homology cloning method and RACE approach. The full length of the cDNA is 1 767 bp long, with a 5′UTR of 38 bp, an ORF of 1 176 bp, encoding 392 amino acids and a 3′UTR of 553 bp. The predicted amino acid sequenceis consisted of a signal peptide of 18 aa, a pro-sequence of 29 aa and a mature protein of 345 aa, with an estimated isoelectric point of 8.04 and molecular mass of 42.3 kDa. The amino acid sequence of its pgCTSDis highly similar with that of Pinctada maxima (79% similarity), and shares 59%~75% similarity with other organisms.The fluorescent quantitative analysis suggests that pgCTSD mRNA expresses in all the tissues tested including adductor muscle, gonad, hepatopancreas, mantle and gill, with higher expression level in hepatopancreas and lower level in adductor muscle. Compared with the control, the expression level ofpgCTSD in the LPS-challenged test groups decreases in tissues of gonad and hepatopancreas significantly but increases in adductor muscle in 6 h after injection of LPS, yet no obvious change was observed in mantle or gill tissues. In 6 h after injection of Vibrio harveyi, the expression profile ofpgCTSD decreases in hepatopancreas and mantle but increases in adductor muscle and gill, yet no significant change is found in gonad. The cDNA expression′s response to challenges of LPS and Vibrio bacteria in hepatopancrea suggests pgCTSD′s participation in immune reaction.
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