Design, construction, sequence analysis and bioinformatics study of RTB-ipaD gene cassette: A new way in generation of Shigellosis vaccine

Shigella is the most common cause of diarrhea. Plasmid antigen ipaD is essential for bacterial invasion into the host cells. One of the challenges of mucosal vaccines against Shigella based on ipaD proteins is its limited potency. It seems that by linking this ipaD to an appropriate carrier and adjuvant such as Ricin toxin B subunit we can render it very immunogenic. This study aimed to construct an expression vector containing RTB-ipaD gene cassette، sequence analysis and bioinformatics study. In this study، RTB and ipaD (163–483) genes were cloned into the pGEM-T vector. ipaD gene was fused to RTB gene with a linker encoding the GPGP in pGEM vector by restriction enzymes method. Then، the fused gene of RTB-ipaD was subcloned into the pET28a (+) expression vector. Finally، sequencing analysis and bioinformatics study of gene cassette was performed. Construct validity of RTB-ipaD gene cassette into the pET28a (+) expression vector confirmed by PCR and enzymatic digestion. The results of sequencing of gene cassette were compatible with the related deposited gene in Genbank. Bioinformatics studies base on the Codon Adaption Index showed that gene cassette is capable of expressing the transgenic plants، and some strains of E. coli. Linking of RTB as a carrier and adjuvant to antigen ipaD is a new and desirable approach in order to generate a mucosal vaccine for shigellosis