Cloning of the Phosphatidylserine Synthase Gene and Its Secretive Expression in Bacillus subtilis / 中国生物工程杂志

ABSTRACT

The gene encoding the phosphatidylserine synthase in Escherichia coli K12 Sgal-(ExPASy P23830) was amplified by PCR. After DNA sequence analysis, it was inserted into the inducible expressive shuttle vector pBES of Bacillus subtilis, which was constructed in the lab, and the recombinant plasmid pBES-pss was transformed into competent cells of the Bacillus subtilis strain DB104. The positive transformant DB104 (pBES-pss) was grown on Bacillus subtilis common fermentation medium, which contained 30?g/ml kanamycin. After 2 hours cultivation, sucrose was added and increased to the final concentration of 2% for induction and this phosphatidylserine synthase was secreted into the medium. The result of SDS-PAGE showed that the molecular weight of the protein was 52kDa and the result of enzyme coupling colorimetric method showed that the enzyme activity was 1.50U/ml. The recombinant Bacillus subtilis has increased the yield of phosphatidylserine synthase which will be used for industrial biosynthesis of phosphatidylserine.