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[Prokaryotic expression and application of SpiC protein of Salmonella pullorum]
Shizhong Geng1, Huan Liu1, Xinan Jiao1
1Jiangsu Provincial Key Laboratory of Zoonoses, Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China.
Objective:
To express the SpiC protein of Salmonella pullorum and establish an indirect ELISA method with SpiC protein as antigen.
Methods:
The 384 bp spiC gene of Salmonella pullorum was amplified by PCR from the genomic DNA and cloned into pET30a vector. The recombinant plasmid pET30a-spiC was transformed into competent E.coli BL21(DE3) cells and induced by IPTG. The expressed product was analyzed by SDS-PAGE and Western blotting. Indirect ELISA based on purified SpiC protein was applied to detect 144 clinical serum samples.
Results:
SDS-PAGE and Western blotting confirmed that a soluble recombinant His-SpiC protein of 19.2 ku was expressed in BL21(DE3) cells. SPF chicken antibodies against GST-SpiC could recognize His-SpiC, indicating that His-SpiC had a good immunogenicity. The indirect ELISA that we established using His-SpiC protein as coating antigen for detecting antibodies against SpiC could differentiate infected from vaccinated animals (DIVA).
Conclusion:
The recombinant His-SpiC was successfully expressed and the indirect ELISA with it as coating antigen could be used as DIVA method for the related vaccine of pullorum disease.
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