2.2. Bacteria culture and genomic DNA extraction
Colonies of E. coli and P. aeruginosa cells were grown on
LB agar plates at 37 °C for 17 h. A single colony was selected from
the plates and inoculated into 5 ml LB broth medium for 16 h. Next,
1-ml culture was serially ten-fold diluted in distilled water and
utilized for colony-forming unit (CFU) assay by the standard
spread-plate technique.
In the genomic DNA extraction protocol, first suspension of cells was
pelleted at 5000 g for 10 min. Next, the pellet was re-suspended in
1 ml of DNAzol® Reagent and incubated for 15 h at room temperature.
Next, the samples were boiled at 10 min, and 500 μl of 100%
Isopropanol were added with vigorous mixing. Then, samples were placed
on ice for 10 min before being centrifuged at 12,000 g for 10 min. The
DNA pellet was washed with 70% (v/v) ethanol, air dried and dissolved
in 300 μl of DNase-free water. Next, we incubated the DNA samples at
65 °C for 10 min. Quality and concentration of the DNA samples were
measured by spectrophotometric analysis at 260 and 280 nm.