Post-Evolution Growth Assays of Sulfolobus acidocaldarius Ancestral and Evolved Lineages

To begin incubate the revived ancestral and temperature evolved cultures of Sulfolobus acidocaldarius at the final temperatures. Once incubation is complete, measure their optical densities at 600 nanometers. Next, add 1.5 milliliters of BBM plus into six sets of seven tubes for each culture.

Then add 15 microliters of each culture into the six sets of tubes. Set three thermomixers to 75 degrees Celsius and three thermomixers to 65 degrees Celsius. Place the tubes in the thermomixer corresponding to the temperature and replicate ID.After a 48-hour incubation transfer, 200 microliters from each culture into a 96-well plate.

Measure the optical density with a spectrophotometer. Finally, plot and analyze the data with statistical software. Lineages from the constant 75 degrees Celsius condition increased in OD600 from an initial range by the end of the experiment.

In contrast, lineages from the constant 65 degree Celsius treatment displayed a drop. Populations in the temperature drop treatment increased from the initial optical density to transfer six, then showed a steady decline.