Posted: May 22nd, 2023
A single colony was taken and inoculated overnight into 2 ml LB/antibiotic overnight, in the shaking incubator at 37oC. The overnight culture from step 1 was added into 10 ml of fresh LB using the most appropriate antibiotic in a 50 ml falcon tube so that the new culture had an OD600Â of 0.1. The contents were placed in the incubator at 37oC and shaken until the OD600Â had reached 0.5-0.8. Once the correct OD600Â had been attained, the value was recorded and 1 ml of culture removed, then placed in an eppendorf tube labeled IPTG.
The tube labeled IPTG was spinned for 5 min at 4000 rpm, and then the supernatant was drained. The pellet was stored in the freezer at -20Â°C to check on the gel. 1mM IPTG was added to the culture that had remained in the falcon tube every 1 hour, 1 ml of culture was then taken, the OD values recorded a new eppi placed, and steps 6-7 repeated once more. In addition, step 8 was repeated for 4 hours and the samples run on SDS-PAGE.
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