Production of Brucella lumazine Synthase Recombinant Protein to Design a Subunit Vaccine against Undulant Fever

Document Type: Original Articles

Authors

1 Department of Animal Science, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran

2 Immunology Research Center, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran

Abstract

Brucella bacterium causes Brucellosis, an infectious disease spreading from animals to human. Brucella lumazine synthase (BLS) is a highly immunogenic protein with adjuvant properties, which has been introduced as an effective protein carrier for vaccine development. This protein also plays a significant role in inducing immune system. This study aimed to clone, express, and purify the BLS gene from Brucella melitensis Rev1. The BLS gene was amplified by particular primers with the restriction enzyme sites as a linker and it was inserted into pTZ57R/T vector. Subsequently, it was ligated into pET32(a)+ expression vector. Recombinant expression vector containing coding sequence of BLS was transformed into E. coli BL21 (DE3) host gene expression and stimulated by 0.1mM IPTG. The results of sequencing showed that there were not any mutations in BLS encoding sequence. The expression results were set by sequencing and endorsed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses and western blotting that showed 35 kDa protein band appropriately.

Keywords

Main Subjects


Article Title [French]

Production de la Protéine Recombinante Brucella lumazine Synthase Afin de Concevoir un Vaccin Sous-unitaire contre la Fièvre Ondulante

Abstract [French]

La bactérie Brucella est à l'origine de la Brucellose, une maladie infectieuse se transmettant de l'animal à l'homme. La Brucella lumazine synthase (BLS) est une protéine hautement immunogène dotée de propriétés adjuvantes, qui a été présentée comme un vecteur efficace de transport de protéines pour le développement de vaccins. Cette protéine joue également un rôle important dans l'induction d’une réponse immunitaire. Cette étude visait à cloner, exprimer et purifier le gène BLS de Brucella melitensis Rev1. Le gène BLS a été amplifié par des amorces spécifiques avec les sites d'enzyme de restriction servant de liant afin d’etre inséré dans le vecteur pTZ57R / T. Ensuite, il a été ligaturé dans le vecteur d'expression pET32(a)+. Le vecteur d'expression recombinant contenant la séquence codante de BLS a été transformé dans le système d’expression génique E. coli BL21 (DE3) et stimulé par 0.1 mM d'IPTG. Les résultats du séquençage ont montré qu'il n'y avait pas de mutations dans la séquence codant pour BLS. L'expression de la protéine recombinante a été vérifiée par séquençage ainsi qu’à l'aide d'analyses électrophorèse sur gel de dodécyl sulfate de sodium-polyacrylamide (SDS-PAGE) et d'un transfert de type western montrant une bande protéique de 35 kDa de manière appropriée.

Keywords [French]

  • Brucella melitensis Rev1
  • BLS
  • Expression génique
  • Protéine recombinante
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