Construction,identification and expression of Enterococcus faecalis based recombinant Efs-Sj14-3-3 vaccine of Schistosoma japonicum
Objective To construct an Enterococcus faecalis(Efs)based recombinant Efs-Sj14-3-3 vaccine of Schistosoma japonicum(Sj)and observe its expression efficiency.Methods The recombinant plasmid pGEX-Sj14-3-3 and competent cells of Enterococcus faecalis ATCC47077 strain were preparated;pGEX-Sj14-3-3 was electroporated into Enterococcus faecalis competent cells to construct recombinant Efs-Sj14-3-3 vaccine;the plasmid was extracted from recombinant Efs-Sj14-3-3 for PCR identification.The recombinant Efs-Sj14-3-3 vaccine was expressed through isopropyl-beta-D-thiogalactopyranoside(IPTG)induction,and the recombinant protein was verified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE)and Western blotting.Results PCR showed that 399 bp Sj14-3-3 gene was amplified when the extracted plasmid from recombinant Efs-Sj14-3-3 as template;the expressed GST-Sj14-3-3 fusion protein was approximately 40×103 Mr by SDS-PAGE;the amount of the expressed protein was 18％of the total bacterial proteins.Western blotting showed that the expressed protein could be recognized by sera from patients infected with Schistosoma japonicum.Conclusion The recombinant Efs-Sj14-3-3 vaccine of Schistosoma japonicum was successfully constructed,and the expressed fusion protein shows specific antigenicity.
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