High-effective expression and construction of a prokaryote expression vector pET30a-glutathione-S-transferase A1 gene from human liver cDNA library and sequence determination

Chen, P.; Zhang, C.; Wu, Y.; Hang, L.; Chen, L.

Wei Sheng Yan Jiu 33(5): 596-599

2004


ISSN/ISBN: 1000-8020
PMID: 15612490
Accession: 049220668

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Abstract
To express highly effective on IPTG-induced and construct an prokaryote expression vector carrying human glutathione-S- transferase (GST) A1 gene and to determine partial sequence analysis. The GSTA1 cDNA was amplified and extracted from human liver total RNAs by RT-PCR approach and recombined with prokaryote expression vector pET30a. The recombined vector pET30a-GSTA1 was verified using PCR, restriction analysis and sequencing determination and induced with IPTG in 35 degrees C . Human GSTA1 gene was recombined corrected with pET30a, compared with Genbank , in code 512 T-->C, amino acid Met-->Thr; alignment core is 99% . The expressed fusion protein,with molecular weight of about 34.4 KDa, were about 41.8% (1h), 68% (2h), 68.3% (3h), 83% (4h) of the total cell protein by SDS-PAGE. The protein expression of GSTA1 was demonstrated by Western blotting, it is correct.