Samples collected from a Midwestern dry-grind fuel ethanol
plant were serially diluted in phosphate buffered saline and spread
on plates containing MRS medium (Difco Becton Dickinson, Sparks,
MD) with 0.001% (w/v) cycloheximide added to suppress yeast
growth (Skinner and Leathers, 2004). Cultures were incubated
anaerobically at 37 C using the Mitsubishi AnaeroPack system
(Thermo Fisher Scientific, Pittsburg PA) or the GasPak EZ System
(Becton Dickinson) and strains were purified twice as single colonies
and stored at 80 C in 40% (v/v) glycerol in 96-well microtiter
plates.
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