Cloning of Metallotionein cDNAs and Its Gene in Shore Crab(Carcinus maenas).
Metallothioneins are cysteine-rich proteins which chelate metal ion(Zn, Cu, Hg and Cd)and regulate ion concentrations in animal body. Based on the published amino acid sequence of C.maenas metallothionein, completely degenerated PCR primers were designed and two different types of cDNA were amplified from gill tissue RNA of C.maenas using RT-PCR method. These cDNAs were cloned to pGEM T Easy Vector and the nucleotic acid sequences were determined. The results showed that one group of the cDNA sequence is exactly same as first 123 bases, as predicted from MTIb cDNA. Another group of cDNA was also amplified, which is truncated and different at the 3' end of cDNA. Based on the former cDNA sequence, specific primers were designed to amplify the full length MTIb cDNA coding region. The MTIb gene was also amplified and cloned into the pGEM Vectors. The gene sequence showed that MTIb gene contains two introns(575 bp and 602 bp), dividing the gene to three exons(22 bp, 77 bp and 78 bp).