Construction of ompW knock-out mutants of Escherichia coli to increase sensitivity to neomycinsulphate and ampicillin

Abstract:

Abstract:［Objective］To investigate the contribution of an outer membrane protein OmpW to tolerance neomycinsulphate and ampicillin of Escherichia coli K12. ［Methods］The ompW knock-out mutant (ΔompW) of E． coli K12 was generated using λ-Red recombination system． Then the minimal inhibitory concentration (MIC) and the survival rates under 1/2 MIC of neomycinsulphate or ampicillin of ΔompW and E． coli K12 were determined respectively． ［Results］The ΔompW was successfully obtained through confirmation of PCR analysis at the gene level and Western blot analysis at the protein level． The MIC of neomycinsulphate of ΔompW is 1.7 μg/mL．The value is much lower than that of E． coli K12，which is 8.0μg/mL． Difference of survival rates under 1 /2 MIC of neomycinsulphate of ΔompW and E． coli K12 was also observed，and their survival rates are 39% and 98% ，respectively． The MIC of ampicillin of ΔompW is 3.3 μg/mL．The value is also lower than that of E． coli K12 (16.0μg/mL)．The survival rates under 1/2 MIC ampicillin of ΔompW and E．coli K12 are 30.3% and 70.38%， respectively．［Conclusion］ The ΔompW is much more sensitive to neomycinsulphate and ampicillin than its parent strain． The result indicated that OmpW played crucial role in bacteria resistance of drug．