Table 3.

Re-introduction of spontaneous adhE and hfs hydrogenase mutations that occurred during growth adaptation of the Δldh , Δpta Δack ethanologen strain M0863 into wildtype (WT) T. saccharolyticum

		Units in mM
Strain	Genotype	Cellobiose consumed	Lactate	Acetate	Ethanol	Carbon recovery (%)
M0010	WT	28.2 ± 0.6	13.8 ± 0.5	34.7 ± 0.7	63.5 ± 2.3	99
M2202	adhE*::kan R	30.2 ± 0.0	2.4 ± 0.1	42.3 ± 0.3	76.5 ± 0.6	100
M2203	adhE::kan R	27.7 ± 2.6	1.4 ± 0.2	29.0 ± 1.6	78.4 ± 8.6	98
M2204	hfs*::kan R	29.9 ± 0.1	20.3 ± 0.5	4.2 ± 0.1	91.7 ± 0.6	97
M2205	hfs::kan R	28.1 ± 0.7	9.9 ± 2.2	29.8 ± 1.1	67.9 ± 2.7	96

*mutations found in evolved strain M0863 introduced by homologous recombination. Data are average of four replicates, anaerobic TSC7 medium, 55°C, 24-h fermentation of 30.4 ± 0.7 mM cellobiose. Mutations were created via homologous recombination of a kanamycin resistance marker downstream of the hfs operon that contained mutations in the upstream flanking region. M2202 - kan^R inserted 3′ of the adhE gene, with the M0863 mutant adhE sequence. M2203 - kan^R inserted 3′ of the adhE gene, with the WT adhE sequence. M2204 - kan^R inserted 3′ of the hfs gene operon, with the M0863 mutant hfs sequence. M2205 - kan^R inserted 3′ of the hfs gene operon, with the WT hfs sequence.