Construction of Recombinant Autolytic E.Coli Cells Producing Bioplastic Via Cloning of Poly(3 Hydroxybutyrate) Operon and Gene E (E.Lysis) Coordinately

Production of material such as synthetic polymers resistant to biodegradation increases wastes and accumulates them in environment for a long time. Hence, the production of biodegradable polymers particularly from renewable and natural resources become necessary. Polyhydroxybutyrate(PHB) is a storage material and biodegradable polymer in a great variety of bacteria. Ralstonia eutropha has been studied extensively because of its ability to accumulate PHB. In this bacteria PHB is synthesized by acetyl coA in three steps by function of three enzymes, β-ketothiolase, NADPH-dependent acetylcoA reductase, PHA synthase which are encoded by phbA, phbB, phbC, respectively. These genes are located in an operon. In this project, A strain of Escherichia coli was engineered to express poly(3 hydroxybutyrate) and gene E (E.lysis) coordinately. A gene encoding E.lysis was placed under the control of the IPTG-inducible T7 promoter in pET28a while the polyhydroxyalkanoate synthesis operon (phaCAB) from R. eutropha was expressed constitutively under the control of its native promoter. After the production of PHB in late logarithmic phase, IPTG was added and gene E expressed. The aim of this project is the construction of autolysing E. coli cells which produce bioplastic.