Isolation of a cDNA Encoding a Chitinase Family Protein from Cuticular Tissues of the Kuruma Prawn Penaeus japonicus

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Abstract
To identify and characterize a chitinase related to molting in the Kurumia prawn Penasus japonicus, we searched for chitinase-encoding cDNAs expressed in cuticular tissues. Using two degenerate oligonucleotide primers derived from the two conserved regions of the chitinase protein family, a RT (reverse transcription)-PCR product was obtained. This product was used as a probe to screen a cDNA library from a mixture of the tall fan and blade-two tissues which consist mainly of chitinous exoskeleton and underlying epidermis. A positive cDNA clone was analyzed for the sequence. This clone contains an open reading frame for a protein (named Pjchi-2) of 527 amino acids which exhibits sequence similarity to known chitinases. A typical signal sequence could not be found in the Pjchi-2 sequence. Significant accumulation of Pjchi-2 mRNA was detected in the mixture of the tall fan and blade prior to molting, whereas the transcript level was much lower during the intermolt stage. This observation suggests that Pjchi-2 plays a role in molting. The mRNA was not detected in the hepatopancreas. This expression pattern of Pjchi-2 makes a contrast to that of Pjchi-1 which encodes another chitinase family protein in P. japonicus, and is expressed in the hepatopancreas but not in the tall fan or blade.