| Polyhydroxybutyrate (PHB) belongs to a class of biodegradable polymers, which has properties similar to conventional plastics. Ralstronia eutropha naturally produces PHB using acetyl-CoA. Acetyl-CoA is also abundantly found in developing seeds of higher plants, where oil synthesis takes place. Three genes responsible for PHB synthesis were isolated from R. eutropha. Two gene constructs were prepared, driven by seed-specific promoters, to drive PHB synthesis in cytoplasm and leucoplast of developing seeds. The cytosolic construct was introduced into Arabidopsis thaliana by Agrobacterium-mediated transformation. Homozygous lines with single copy transgenes were developed and characterized at molecular level. Transgenic seeds were analyzed by Gas Chromatography and found to accumulate 0.772 +/- 0.006% PHB (dry weight). Furthermore, phb genes were over-expressed in E. coli and recombinant Phb proteins were purified. Successfully purified Phb proteins were used for immunization into rabbit hosts to produce polyclonal antibodies. |
