Antibiotic susceptibility testing is the most suitable strategy for antibiotic resistance detection in bacteria. This project was an initial step for the production of β-lactamase for clinical diagnostics in Pakistan. The current work deals with the optimization of conditions for higher level production of recombinant β-lactamase from Bacillus subtilis R5. Supplementation of LB medium with various carbon sources including wheat bran, rice bran and molasses could increase the enzyme production from 25 to 43.7, 44.3 and 49 µmole/min whereas the nitrogen sources including tryptone, peptone and yeast extract could enhance the enzyme production from 25 to 40.21, 41 and 43.76 µmole/min, respectively. The highest β-lactamase production was recorded when the LB medium was supplemented with 4% wheat bran and 2% yeast extract. The locally produced recombinant β-lactamase exhibited strong potential regarding the hydrolysis of β-lactam ring containing antibiotics and showed comparable results to β-lactamase from Hi-media kit available in the market. The β-lactamase from current study was found suitable for its use as positive control in antibiotic susceptibility testing and this enzyme will be utilized for the development of antibiotic susceptibility testing kit at domestic level in near future.
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