PlpE Epitopes of Pasteurella multocida Fusion Protein as Novel Subunit Vaccine Candidates
| Autor: | Gholamreza Nikbakht Brujeni, Mohsen Abolhassani, Soroush Sardari, Saied Mostaan, Mohammad Ali Shokrgozar, Abbas Ghasemzadeh, Parastoo Ehsani |
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| Přispěvatelé: | Molecular Biology Department [Tehran, Iran], Institut Pasteur d'Iran, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Biotechnology Research Center, National Cell Bank of Iran [Tehran, Iran] (NCBI), Department of Immunology [Tehran, Iran], University of Tehran, This project was honorably sponsored by the Pasteur Institute of Iran as Ph.D. dissertation project. |
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
vaccine candidate [SDV]Life Sciences [q-bio] 030106 microbiology Virulence lcsh:Medicine Biology medicine.disease_cause Epitope Microbiology law.invention 03 medical and health sciences 0302 clinical medicine law medicine 030212 general & internal medicine Pasteurella multocida Pasteurella Multocida Escherichia coli lcsh:QH301-705.5 Polymerase chain reaction Fusion PlpE Cloning Immunogenicity lcsh:R General Medicine biology.organism_classification Fusion protein 3. Good health lcsh:Biology (General) Original Article |
| Zdroj: | Advanced Biomedical Research Advanced Biomedical Research, Wolters Kluwer, 2020, 9 (1), pp.43. ⟨10.4103/abr.abr_245_19:10.4103/abr.abr_245_19⟩ Advanced Biomedical Research, Vol 9, Iss 1, Pp 43-43 (2020) |
| ISSN: | 2277-9175 |
| Popis: | International audience; Background: Pasteurella multocida is the causative agent of many diseases. Antimicrobial treatment disadvantages highlight the need to find other possible ways such as prophylaxis to manage infections. Current vaccines against this agent include inactivated bacteria, liveattenuated bacteria, and nonpathogenic bacteria, which have disadvantages such as lack of immunogenicity, reactogenicity, or 1 1 2 3 4 1 2 3 4 reversion to virulence wild bacteria. Using bioinformatical approaches, potentially immunogenic and protective epitopes identified and merged to design the best epitope fusion form in case of immunogenicity as a vaccine candidate. Materials and Methods: In this study, the fusion protein (PlpE1 + 2 + 3) and full PlpE genes (PlpE-Total) were cloned in pET28a in BL21 (DE3) firstly and later in pBAD/gIII A and expressed in Top10 Escherichia coli. Overlap polymerase chain reaction (PCR) using different primers for 5ˈ and 3ˈ end of each segment produced fusion segment 1 + 2 and (1 + 2) +3 fragments and was used for cloning. Results: Cloning of both PlpE1 + 2 + 3 and PlpE-Total into the pET28a vector and their transform into the BL21 (DE3) E. coli host was successful, as the presence of the cassettes was proved by digestion and colony PCR, however, their expression faced some challenges independent of expression inducer (isopropyl β-d-1-thiogalactopyranoside) concentration. Conclusion: Changing the vector to pBAD/gIII A and consequently changing the host to Top10 E. coli have resulted in sufficient expression, which shows that Top10 E. coli may be a good substitute for such cases. Furthermore, it is concluded that adding 8M urea results in sufficient purification, which hypothesizes that denature purification is better for such cases than native one. Purified proteins headed for further analysis as vaccine candidates. |
| Databáze: | OpenAIRE |
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