In Vitro And Vivo Studies Ofchicken Egg Yolk Antibody (IgY) Against Helicobacter Pylori

PARTâ… Cloning and Expression of NapA-ctxB fusion protein to Helicobacter pyloriObjective: To construct and express the fusion gene of H.pylori NapA and cholera toxin subunit B(ctxB) by gene-engineering, which would lay a foundation for the development d vaccine for H.pylori infection.Methods: A recombinant strain which could express bivalent antigen of NapA and CtxB subunit was constructed. CtxB gene was amplified by PCR and cloned into plasmid pQE30-napA. The recombinant plamsid was identified by sequencing ,then transformed into E.coli DH5a.. The transformant colony was induced with IPTG. Purify the expressed protein by Ni2+-NTA column chromatography. Expression of fusion protein was analyzed by SDS-PAGE and Western blot.Results: The gene fragment at length of 807 bp was amplified and successfully cloned into plasmid pQE30.SDS-PAGE showed a protein band with relative molecular weight of 30 000,which was consistent with the expectation.The expressed product contained about 27% of total somatic protein ,and reached a purity of 94% after Ni2+-NTA column chromatograph.Western blot showed good antigenicity of the recombinant protein with the serum of anti-NapA and anti-CT.Conclusion: The recombinant plasmid of neutrophil activating protein(NapA)and cholera toxin subunit B(CtxB) was successfully constructed and NapA-CtxB gene was highly expressed in E.coli DH5a.