ISOLATION AND CLONING OF cDNA OF GENE ENCODING FOR METALLOTHIONEIN TYPE 2 FROM MELASTOMA AFFINE

Authors

  • SUHARSONO SUHARSONO 1 Research Center for Bioresources and Biotechnology
  • NIKEN TRISNANINGRUM 1Research Center for Bioresources and Biotechnology
  • LULUT DWI SULISTYANINGSIH 1Research Center for Bioresources and Biotechnology
  • UTUT WIDYASTUTI 1Research Center for Bioresources and Biotechnology, and 2Department of Biology, Bogor Agricultural University Indonesia

DOI:

https://doi.org/10.11598/btb.2009.16.1.64

Abstract

Metallothionein is an important protein for detoxifying heavy metal ions. This research was conducted to isolate and clone cDNA of gene encoding for metallothionein type 2 from Melastoma affine. Total RNA was isolated from young leaves. Total cDNA was synthesized from the total RNA by reverse transcription. The MaMt2 cDNA was successfully isolated by PCR technique. The MaMt2 cDNA was inserted into pGEM-T Easy and the recombinant plasmid was successfully introduced into Escherichia coli DH5α. DNA sequencing analysis showed that this cDNA is full length consisting of 246 pb encoding 81 amino acid residues. This cDNA is identical to mRNA of AtMt2 from Arabidopsis thaliana. It does not contain any restriction sites found in the cloning sites of pGEM-T easy. The deduced protein of MaMT2 contains 14 cysteine residues distributed in the Cys-Cys, Cys-X-Cys, and Cys-X-X-Cys motifs.   Key words: cDNA, metallothionein, Melastoma affine, cloning, cysteine

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