MEC136 strain evolution with acetate as the sole carbon source (parent is ATCC 8739 E coli)
Adaptive evolution using E. coli ATCC 8739 was conducted in a chemostat with 800 mL defined medium in a 2.5 L bioreactor (Bioflo 2000, New Brunswick Scientific Co. Edison, NJ, USA). Air was sparged into the bioreactor at a flow rate of 1.0 L/min, the agitation was 500 rpm, the temperature was maintained at 37°C, and the pH controlled at 7.0 using 20 % (w/v) NaOH and 20% (v/v) H2SO4.  The feed used for adaptive evolution contained 35 mM acetate, and all other nutrients at one-quarter the concentrations used in initial shake flask studies (1.70 g citric acid, 13.30 g KH2PO4, 4.50 g (NH4)2HPO4, 1.2 g MgSO4·7H2O, 13 mg Zn(CH3COO)2·2H2O, 1.5 mg CuCl2·2H2O, 15 mg MnCl2·4H2O, 2.5 mg CoCl2·6H2O, 3.0 mg H3BO3, 2.5 mg Na2MoO4·2H2O, 100 mg Fe(III) citrate, 4.5 mg thiamine·HCl, 8.4 mg Na2(EDTA)·2H2O).  When the dilution rate reached 0.50 h-1, the culture was plated onto LB plates, MEC136 were selected for further study and sequencing.  MISeq using the MiSeq Reagent Kit v3 (600 cycles), catalog number MS-102-3003 was performed on DNA isolated from the parent and evolved strains by Research and Testing Laboratory (Lubbock, TX).