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DNA extraction and PCR screening using primers designed on conserved region of PGA genes. The PCR product from a positive isolate were cloned and subjected to sequencing. The gene encoding for full length PGA was expressed in E. coli under the T7 promoter. PCR screening identified several PGA positive E. coli. One of the positive isolates was cloned in pGEM -T easy vector. Sequencing of the cloned gene revealed that the gene encoding Penicillin G acylase from this wild E. coli isolate contains
an open reading frame of 2538 nucleotide encoding 846 amino acids. Analysis of the sequencing results showed that the PGA gene is highly conserved among E. coli strains. Recombinant expression of PGA from wild isolate in E. coli BL 21 resulted in a high level expression of recombinant PGA which appeared as a dense band in SDS-PAGE analysis of induced culture.