Computational protocol: Immunogenicity of Multi-Epitope DNA and Peptide Vaccine Candidates Based on Core, E2, NS3 and NS5B HCV Epitopes in BALB/c Mice

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Protocol publication

[…] An arrangement of the selected epitopes with suitable spacer combination was designed using two computer based modeling for peptide binding (http://imed.med.ucm.es/Tools/rankpep.html) and optimization of proteasomal processing (http://www.paproc.de/). Furthermore, to minimize creation of junctional epitopes, a suitable arrangement of epitopes was designed using two web-based algorithms of SYFPEITI and BIMAS.According to mammalian Mus musculus codon frequency table (www .kasusa.org.jp/codon), the nucleotide sequence of the construct was further optimized to support mammalian codon usage. The Kozak sequence was added to the N-terminus of the multi-epitope construct to enhance its expression. The multi-epitope DNA (PT) was designed with suitable restriction enzyme sites (HindIII and BamHI) at the 5΄ and 3΄ ends, respectively. The PT construct was synthesized by Biamatik (Biomatik Corporation, Canada).Constructed HCV DNA PT was digested with HindIII and BamHI restriction enzymes and inserted to the corresponding sites of expression vectors, pEGFP-N1 and pcDNA3.1 (+) (Invitrogen, Germany). The plasmid with the correct insert was detected by restriction analysis, PCR and sequencing.pEGFP-N1 plasmid containing PT (pEGFP-PT) was purified by an alkaline lysis method (Qiagen, Plasmid Midi kit, Germany). The pcDNA-PT and pcDNA3.1 (+) plasmids were purified by ion exchange chromatography with EndoFree plasmid Purification Giga kit (Qiagen, Germany). DNA concentration was determined by analyzing the absorbance at 260 nm. […]

Pipeline specifications

Software tools RANKPEP, PAProC
Application Immune system analysis
Organisms Classical swine fever virus, Mus musculus
Diseases Infection