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Expression cassette and plasmid construction for Yeast Surface Display in Saccharomyces cerevisiae |
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| Authors: | Piraine Renan Eugênio Araujo Gonçalves Vitória Sequeira dos Santos Junior Alceu Gonçalves Cunha Rodrigo Casquero de Albuquerque Pedro Machado Medeiros Conrad Neida Lucia Leite Fábio Pereira Leivas |
| Institution: | 1.Núcleo de Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brazil ;2.Programa de Pós-Gradua??o em Veterinária, Faculdade de Veterinária, Universidade Federal de Pelotas, Pelotas, RS, Brazil ; |
| Abstract: | Objectives Develop a Cell Surface Display system in Saccharomyces cerevisiae, based on the construction of an expression cassette for pYES2 plasmid. ResultsThe construction of an expression cassette containing the α-factor signal peptide and the C-terminal portion of the α-agglutinin protein was made and its sequence inserted into a plasmid named pYES2/gDαAgglutinin. The construction allows surface display of bovine herpesvirus type 5 (BoHV-5) glycoprotein D (gD) on S. cerevisiae BY4741 strain. Recombinant protein expression was confirmed by dot blot, and indirect immunofluorescence using monoclonal anti-histidine antibodies and polyclonal antibodies from mice experimentally vaccinated with a recombinant gD. ConclusionsThese results demonstrate that the approach and plasmid used represent not only an effective system for immobilizing proteins on the yeast cell surface, as well as a platform for immunobiologicals development. |
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