After allowing the library three days to repopulate, we inoculated 50 μL of the selection culture into 2.5 mL fresh LB/carb in preparation for a second round of selection. The outgrowth was subsequently grown to an OD of 0.5, then a 100 μL aliquot was inoculated into 2.5 mL LB/carb containing 50 mM selenite and 50 uM IPTG. The 50 mM selenite concentration was maintained in the second round to directly compare growth rates and assess if selection had occurred in the first round -under the hypothesis that a fitter population would better withstand an equal challenge.