COLONING AND EXPRESSION OF FLIC AS A VACCINE CANDIDATE AGAINST TYPHOID

Salmonella enterica is a zoonotic pathogen causing typhoid fever in humans and
animals. It is an important cause of food borne infections in humans throughout the world. A recent
study estimated approximately 22 million cases of typhoid each year with at least 200,000 deaths. FliC
encoding flagellin plays a role in pathogenesis and is an important antigen in vaccination. In this study,
recombinant protein fliC as a candidate vaccine against salmonellosis was produced and purified.
Materials & Methods: The protein sequence of fliC was obtained from NCBI database. For high level
expression of protein, the gene was synthesized with codon bias of E. coli.; synthetic gene in pUC57
was sub coloned into pET32 expression vector and then transferred into E. coli BL21DE3. The
recombinant protein was express by IPTG induction. For high expression, three parameters including
IPTG concentration, time and temperature of induction were optimized. The recombinant protein was
purified by ion exchange chromatography with His-Tag. The purified protein was confirmed by western
blotting.
Results: Recombinant vector was approved by PCR and restriction analysis. SDS-PAGE showed a 51
kDa protein with proper purity and western blotting with his-tag antibody confirmed the intend protein.
Conclusion: The use of recombinant vaccines is highly regarded, since one of the major virulence
factors of Salmonella gene was cloned and the protein was produced and purified in this study. This
protein can be used in candidate vaccine against typhoid