Rapid DNA extraction of bacterial genome using laundry detergents and assessment of the efficiency of DNA in downstream process using polymerase chain reaction

Genomic DNA extraction from bacterial cells involves processes normally performed in most biological laboratories. Therefore, various methods have been offered, manually and kit, but these methods may be time consuming and costly. In this paper, genomic DNA extraction of Pseudomonas aeruginosa was investigated using some laundry detergent brands available in Iran. Afterwards, efficiency of the detergents was compared with manually standard methods and kits. To evaluate the efficiency of the genomic DNA in the processes in which DNA is used as a template, the polymerase chain reaction (PCR) tests and enzyme digestion of PCR product were used. The results show that the detergents could be used to extract genomic DNA. Among the brands studied, five-enzyme Taj and three-enzyme Saftlan had the best performance compared to standard methods.Key words: Bacterial genome, DNA extraction, laundry powder, detergent

The Cytoplasmic and Periplasmic Expression Levels and Folding of Organophosphorus Hydrolase Enzyme in Escherichia coli

BACKGROUND: Organophosphorus hydrolase (OPH) is a type of organophosphate-degrading enzyme which is widely used in the bioremediation process. OBJECTIVES: In this study, the periplasmic and cytoplasmic productions and the activity of recombinant OPH in Escherichia coli were investigated and compared using two pET systems (pET21a and pET26b). MATERIALS AND METHODS: The sequence encoding the opd gene was synthesized and expressed in the form of inclusion body using pET21a-opd and in the periplasmic space in pET26b-opd. RESULTS: Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed a band of about 37 kDa with a maximum expression level at 30°C from pET21a-opd.However, the obtained results of the periplasmic space extraction of OPH (pET26b-opd) showed a very weak band, while the cytoplasmic expression of OPH (pET21a-opd) produced a strong protein band. CONCLUSIONS: The activities studied by the production of PNP were determined by following the increase at 410 nm. The maximum PNP was produced at 30°C with an optical density of 10.62 in the presence of cytoplasmic expression of OPH (pET21a-opd). Consequently, our results suggest cytoplasmic expression system as an appropriate candidate with a high amount of OPH in spite of inclusion body formation, which needs an additional refolding step