[Construction and expression of recombinant plasmid pET28alpha-Sj26GST of Schistosoma japonicum in Escherichia coli BL21].

OBJECTIVE

To construct and express recombinant plasmid pET28a-Sj26GST of Schistosoma japonicum (Sj) in Escherichia coli BL21 (DE3).

METHODS

The total RNA was extracted from Sj adult worms by ultrasound-breaking. The Sj26GST antigen gene was amplified by RT-PCR from the total RNA, and then cloned into prokaryotic expression plasmid pET28alpha and transformed into E. coli BL2 (DE3). The BL21(pET28a-Sj26GST) was induced with isopropyl-beta-D-thiogalactopyranosid (IPTG), and the expressed products were analyzed and identified with SDS-PAGE and Western blot.

RESULTS

The 676 bp Sj26GST gene was successfully amplified by RT-PCR and cloned into pET28alpha. The recombinant plasmid pET28a-Sj26GST was successfully constructed, with a relative molecular weight of expressed recombinant protein at approximately 36 x 10(3) as determined by SDS-PAGE. The amount of the expressed protein comprised 26% of the total bacterial proteins. The fusion protein could be recognized by the sera of rabbits infected with Sj.

CONCLUSION

The recombinant plasmid pET28alpha-Sj26GST is successfully constructed and highly expressed in E. coli in a fused form with His-tag. The expressed fusion protein shows specific antigenicity.