Cloning and sequencing of phbA gene of poly-beta-hydroxybuty rate synthesis and its expression analysis

The gene of the first key enzyme of poly-beta- hydroxybutyrate synthesis, 3 -ketothiolase, has been amplified and cloned from chromosomal DNA of Alcali genes eutrophus H16 by PCR. DNA sequencing results show that phbA cloned in pBluescriptSK(+) has an identical sequence with that reported previously e xcept for one base pair. The plant constitutive expression vector has been constructed and tobacco has been transformed in order to examine the phbA g ene function and the efficiency of ctp gene product, SDS-polyacrylamide gel electrophoresis result shows that the ctp gene product could direct foreig n protein into plastid efficiently and phbA gene could be translated into c orresponding protein with correct size. The enzyme activity analysis of 3-k etothiolase shows that the enzyme could catalyze acetyl-CoA to form acetoac etyl-CoA.