Figure 1

The metabolic pathways for the production of L-lactate by engineered E. coli from glucose. Relevant genes are shown. pps, PEP synthase; pflB, pyruvate formatelyase; ldhA, fermentative D-lactate dehydrogenase; dld, D-lactate dehydrogenase; poxB, pyruvate oxidase; pta, phosphotransacetylase; ackA, acetate kinase; adhE, alcohol dehydrogenase; frdA, fumaratereductase; lldD, FMN-dependent L-lactate dehydrogenase; ldh _Lca, L-lactate dehydrogenase from Lactobacillus casei CICIM-CU B1192; ldh _Strb, L-lactate dehydrogenase from Streptococcus bovis CGMCC 1.1624; ldh _Bcoa, L-lactate dehydrogenase from Bacillus coagulans CICIM B1821. Box A presented the homologous recombination on the chromosome in B0013-090B series. The deletion/expression cassette was first electroporated into E. coli B0013-080C. The transformants were selected within LB medium containing gentamicin and the correct double replacement recombinants were confirmed by PCR. The second selection was carried out by incubating the correct recombinants in LB medium without gentamicin. The correct recombinants were confirmed by PCR. Box B outlined the construction procedure for B0013-090B1, and B0013-090B2 and B0013-090B3. P _lacA -ldh* represented one expression cassette of ldh _Lca, ldh _Strb and ldh _Bcoa under the control the promoter of E. coli ldhA. Abbreviations: PEP, phosphoenolpyruvate; Ac-CoA, acetyl-CoA; Cit, citrate; IsoCit, isocitrate; α-KG, α-ketoglutarate; Fum, fumarate; Mal, malate; OAA, oxaloacetate; GLY, glyoxylate.