Construction, expression of truncated pET41a-Em 18 prokaryotic plasmid and its primary antigenic analysis

Objective: To clone, construct and express truncated pET41a-Em18. 1 recombinant plasmid and to study its primary antigenicity. Methods: The primers of truncated Em18 were designed by DNAman biosoftware and the truncated fragment was amplified by PCR from pMD18-T/Em18, and was cloned into prokaryotic expression plasmid pET41a to construct the pET41a-Em18-1. The recombinant plasmid was analyzed by sequenceing. The rEm18.1-GST fusion protein was expressed by induction with IPTG and purified using His-tag column, and then was detected by SDS-PAGE and Western Blot. Results: The pET41a-Em18-1 positive clone was the exact recombinant plasmid and the expressed rEm18.1-GST recombinant protein can be detected as a band of 41KDa by SDS-PAGE and Western blot which confirmed that the recombinant protein could specifically react with the serum samples from patients with alveolar echino-coccosis (AE). Conclusion:The truncated pET41a-Em18.1 was constructed successfully and the rEm18.1-GST recombinant protein is expressed and shows a good antigenicity. This work has potential for use in the research of epitope analysis of Eml8 antigen.