Abstract
Most cellular processes are mediated by multi-subunit protein complexes which have attracted major interest in both academia and industry. Recombinant production of such entities in quantity and quality sufficient for functional and structural investigations may be extremely challenging and necessitate specific technologies. The baculovirus expression vector system is widely used for the production of eukaryotic multiprotein complexes, and a variety of strategies are available to assemble transfer vectors for the generation of recombinant baculoviruses. Here we detail applications of homology-based cloning techniques for one-step construction of dual promoter baculovirus transfer plasmids and of restriction-free (RF) cloning for the modification of existing constructs.
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Acknowledgments
This work was supported by the Centre National pour la Recherche Scientifique (CNRS), the Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Strasbourg (UdS), the Association pour la Recherche sur le Cancer (ARC), the Ligue nationale contre le cancer, the Institut National du Cancer (INCa; INCA 9378), and the Agence National pour la Recherche (ANR-12-BSV8-0015-01 and ANR-10-LABX-0030-INRT under the program Investissements d’Avenir ANR-10-IDEX-0002-02). Instruct (R&D Project Funding), Instruct-ULTRA as part of the European Union’s Horizon 2020 (Grant ID 731005), and Instruct-ERIC are also acknowledged.
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Rossolillo, P., Kolesnikova, O., Essabri, K., Zamorano, G.R., Poterszman, A. (2021). Production of Multiprotein Complexes Using the Baculovirus Expression System: Homology-Based and Restriction-Free Cloning Strategies for Construct Design. In: Poterszman, A. (eds) Multiprotein Complexes. Methods in Molecular Biology, vol 2247. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1126-5_2
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DOI: https://doi.org/10.1007/978-1-0716-1126-5_2
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