Team:UCLA/Notebook/Recombinant Expression/2 July 2015

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Start-Up Culture Preparation, continued

The three start-up cultures are removed from the shaking incubator. An Optical Density (OD) check was performed on each one.

Start-Up Culture from G-Block 3: 0.2

Start-Up Culture from G-Block 4, Colony 1: 0.38

Start-Up Culture from G-Block 4, Colony 2: 0.28

Procedure to Measure OD

1 mL of LB broth in cuvette used as blank

100 ul of sample, 900 ul of LB broth in cuvette

AD 600 reading multiplied by 10 to obtain OD measurement (Beer-Lambert's Law)

Next, we autoclaved 1 L LB, added 1 mL of Ampicillin. The OD measurement from start-up culture tube labelled G-Block 4, Colony 1, was 0.38, the highest and most optimal out of the three. The contents of this tube were added to the 1 L of LB and 1 mL of ampicillin. The contents were put in a shaking incubator for 3 hours and 45 minutes. Another OD check was completed, this time reported to be 0.5. Next 5 mL of IPTG was added to further initiate growth and the contents were left to incubate overnight.