Figure 1
Design of a dual expression cassette for SaoA and Eno and phenotypic identification. A Schematic diagram of operon fusion of saoA and eno dual expression cassette. A ribosomal binding site was added between the open reading frames of SaoA and Eno, and the nucleotide sequence encoding OmpA secretion signal peptide was fused with the open reading frame of Eno. Both saoA and eno are regulated by the Ptrc promoter. B Plasmid maps of control vector plasmid pYA3493 and pS-SE (SaoA-Eno). C Regulated decreased synthesis of LacI and regulated delayed synthesis of SaoA or Eno proteins in rSC0016(pS-SE) containing ΔrelA::araC PBAD lacI TT mutation. Strain rSC0016(pS-SE) was grown in NB with L-arabinose and D-mannose (Lane 1) and then diluted 1:10 into fresh NB without L-arabinose and D-mannose until OD600 to 0.8. The process was repeated four times (Lane 2–5). Each lane was loaded with around 2.5 × 107 CFU bacteria. Synthesis of LacI, SaoA, Eno and GroEL were detected by western blot using correspondent antibodies. D Growth of rSC0016(pYA3493), rSC0016(pS-SaoA), rSC0016(pS-Eno), and rSC0016(pS-SE) in Nutrient broth with 0.2% L-arabinose and no 0.2% D-mannose. (ns) means that there is no significant difference between rSC0016(pS-SE) and rSC0016(pS-SaoA) or rSC0016(pS-Eno).