Construction of a genetic cassette to express immunogenic dengue virus protein ina plantsystem.

MARTÍNEZ CA,; GIULIETTI AM; RODRÍGUEZ TALOU J.

Mar del Plata, Argentina

Congreso; XLIII Congreso de SAIB, Noviembre 2007, .; 2007

SAIB

Production of recombinant proteins in plant systems has received great attention because of the advantages compared with others production systems. Plant cells can glycosilate and do the posttranscriptional arrangements need for complex glycoproteins. Dengue virus genome encodes three structural (C, prM, E) and seven non-structural proteins. E is an immunogenic glycoprotein and needs to be directed to the secretory pathway through the Nterminus signal peptide (SP), to be N-glycosilated. The presence of an ER retention signal in the C-terminus (KDEL) of some proteins increases their stability and yield in plant systems. The aim of this work is to obtain a genetic construct carrying E and CprME protein with a SP and KDEL cloned in a binary plasmid. E protein was amplified from plasmid pMT/V5-HisA given by Dr. Gamarnik and PS was obtained from pCAMBIA1305.2 plasmid by PCR. CprME was obtained from pMT vector by enzyme restriction. Promoter (35S) and terminator (T) were obtained by PCR from pMOG843 plasmid. The intermediate constructs were cloned into pGEM T Vector and sequenced.We obtained the genetic constructions: 35SPS- E-KDEL-T, 35S-PS-CprME-KDEL-T, 35S-PS-CprME-T and 35S-GRP-E-Term. We cloned these constructions into a pCAMBIA1305.2 binary vector and we are currently doing the Agrobacterium-mediated plant transformation.