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Egypt. J. Exp. Biol. (Bot.). 2012; 8(1): 43-49


MOLECULAR CLONING, SEQUENCE ANALYSIS AND EXPRESSION OF THE GLYCYL-TRNA SYNTHETASE GENE (GLYS A) FROM BACILLUS SUBTILIS SADATA-73

Atef M. Ibrahim.

Abstract
The glycyl-tRNA synthetase (β-subunit) gene from locally isolated and identified Bacillus subtilis sadata-73 was cloned in lambda ZAP XR genomic library XhoI and EcoRI digest. The complete open reading frame was obtained in two steps, sequenced and submitted in the GenBank under accession number EU056819.The gene for this aminoacyl-tRNA synthetase has an open reading frame of 1479 nucleotides. The supposed amino acid sequence encodes a protein of 492 amino acids with MW = 55,142. The protein sequence has extensive overall identity / similarity with the Bacillus subtilis subtilis 168, the Bacillus amyloliquefaciencs, Bacillus licheniformis ATCC 14580 and Bacillus pumilus glycyl-tRNA synthetases (98%, 98%, 78%, 69%, and 66%, respectively). The enzyme was overexpressed as a fusion protein in E.coli using pMal-c2 expression system containing the T7 RNA polymerase/ promoter. The predicted MW for the MBP gene product is in good harmony with the size of the fusion protein determined by SDS-PAGE (M(r) = 55) and the predicted protein has an isoelectric point 5.08.

Key words: glycyl-tRNA synthetase gene, Bacillus subtilis sadata-73



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