[Construction and prokaryotic expression of recombinant gene c-Abeta-c and the immunogenicity analysis of the fusion protein].
2005
AIM: To construct the recombinant prokaryotic expression plasmid pHGhis/c-Aβ-c and evaluate the immunogenicity of the fusion protein expressed in E.coli DH5α. METHODS: The gene fragments of HBc1-71, HBc88-144 and Aβ_1-42 were amplified by PCR. Then the Aβ_1-42 gene was inserted between HBc1-71 and HBc88-144, yielding the recombinant gene c-Aβ-c. c-Aβ-c gene was cloned into pGEMEX and then subcloned into pHGhis plasmids. c-Aβ-c fusion protein expression in transformed E.coli DH5α was induced at 42℃. The expressed fusion protein was analyzed by SDS-PAGE. Six BALB/c mice recieved intraperitoneal injection (i.p) of c-Aβ-c fusion protein purified by saturated ammonium sulfate. The anti-Aβ antibody was detected by indirect ELISA. RESULTS: The recombinant gene was confirmed by restriction enzyme digestion and DNA sequencing. After temperature induction, fusion protein was expressed and existed in the sediment of the bacterial lysate. The expression level was 16% of all the proteins in the sediment. After 3 times of immunization, the titer of anti-Aβ antibody in sera of BALB/c mice reached up to 1∶16 000, while the anti-HBc antibody was undetectable. CONCLUSION: Recombinant c-Aβ-c gene can be expressed in E.coli DH5α. The expressed protein exists in the sediment of the bacterial lysate and has a strong immunogenicity. This study lays the foundation for the experimental animal study of Alzheimer's disease genetic engineering vaccine.
Keywords:
- Correction
- Source
- Cite
- Save
- Machine Reading By IdeaReader
0
References
0
Citations
NaN
KQI