Adaptation of Recombinant Escherichia Coli LSBJ for PHA Production in a Bioreactor

Polyhydroxyalkanoates (PHAs) are plastic biopolymers produced by bacteria under stressful conditions. PHAs are biodegradable and can be made to exhibit similar properties to petroleum-based plastics, which makes mass production of them highly desirable. In this study, we adapted E. coli LSBJ for high biomass production in a 2.2 L New Brunswick Scientific Bio Flo 310 bioreactor and began studies toward high production of PHA polymer in this setting. With the modified ~-oxidation pathway of the LSBJ strain of E. coli, the repeating unit length and resulting properties of the PHA polymer can be controlled. The LSBJ strain was modified with several genetic manipulations (!),aas, !),arcA, !),did, crp*) to assist in future polymer production in the bioreactor, and a method for high biomass production of the strain in this setting was developed. This method has consistently achieved high cell densities, with optical densities at 600 nm in the range of 30-50 and as high as 74, similar to what has been previously demonstrated for E. coli fermentations. This method has also consistently produced biomass several fold higher than shake flask procedures with the LSBJ strain, as evidenced by cell dry weights. With this method fulJy developed for the 2.2 L bioreactor, PHA production methods at high biomass in this vessel can be investigated beyond the preliminary studies already conducted. Scale-up procedures to larger vessel volumes can then be started, beginning the process toward mass production of PHA using the LSBJ strain