Subcloning of DNaseB Gene from Group A of Streptococcus and Constructing pET-28a(+)-DNaseB Plasmid

A 0.5 kb dock-tailed DNaseB gene fragment was amplified by PCR from plasmid pMD18-T-DNaseB.In order to identify its correctness and reproduce this gene abundantly.First,the target gene fragment was cloned into pMD19-T.Then the DNaseB gene was subcloned into pET-28a(+) resulting pET-28a(+)-DNaseB.The recombinant plasmid was transformed into E.coli DH5a.The recombinant plasmid DNA was extracted by SDS Alkaline Lysis Method and purified,and was studied in detail by restriction endonuclease and nueleotide sequencing.The result showed that the nucleotide sequence of the inserted DNA fragment was identical to that of DNaseB gene.The recombinant plasmid was transformed into E.coli BL21(DE3).The target protein was expressed by IPTG induction.