Microbiology
Chemical transformation of yeast requires the organism to be in logarithmic growth phase, so, before we clone our casein genes into yeast, we need to make sure that we know how to grow our yeast to log Phase.
Here's a brief protocol for monitoring yeast growth.
- GROWTH CURVE
(1) Figure out growth curve of yeast strains (we have Carolina strain now, we want ATCC strain, and Keoni Strain)
(a) please be conservative with YPD media here - we can do these experiments in 100mL tubes - in triplicate. Record the OD (650nm) at 0, 5, 10, 24, 48, and 72 hours.
(2) Repeat growth curve with pD1214:FAKS-hKappa cloned into yeast strain.
(a) no, it's not the real human kappa casein, but it'll get us closer to understanding how our strains will perform after real cloning happens. This is a great set of experiments for folks who've not done cloning before. Bacterial transformation and plating, Plasmid Prep, Yeast transformation.
(b) grow the yeast clones in triplicate (100mL) to create growth curve.
So, I think the way to do these growth curves, is to grow an overnight 5mL YPD culture in a 15mL tube from a single yeast colony, read the OD the next morning (>16hrs later, should be around 1.8?), then start the culture with a KNOWN amount of 16-hr innoculum. Record the OD (650nm) at 0, 5, 10, 24, 48, and 72 hours.