Background: Escherichia coli
does not produce n-butanol naturally, but can be butanologenic when related
enzymes were expressed using inducible elements on plasmids. In this study we attempted to confer
E. coli
strain capability of automatic excretion of the chemical by employing a native anaerobic promoter. Also, a
novel DNA kit was designed for PCR preparation of linear DNA fragments to perform strain modification. The
kit is primarily composed of two mother vectors, co-transformation of linear DNAs into
E. coli can
simultaneously introduce two butanol synthetic operons into the chromosome and create two in-frame gene
deletions at targeted native loci.
Results: E. coli strain Bw2V carries plasmid pCNA-PHC and pENA-TA, both utilizes native anaerobic promoter P
hya
for the expression of butanol synthetic enzymes. When Bw2V was subjected in anaerobic fermentation using
medium containing extra glucose, the accumulated n-butanol in the broth was up to 2.8 g/L in bioreactor; as
the genetic element expressing the same pathway was introduced into the genome, the titer of butanol was
1.4 g/L.
Conclusions: The expression systemusing P
hya is effective in applications that involve expression plasmids as also
applicable in ectopic expression as single copy on the chromosome. Results imply that P
hya can be subjected for
broader application in bioproduction of more feedstock chemicals.
