FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi
Fungal synthetic biology is a rapidly expanding field that aims to optimize the biotechnological exploitation of fungi through the generation of standard, ready-to-use genetic elements, and universal syntax and rules for contributory use by the fungal research community. Recently, an increasing numb...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-08-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1222812/full |
| _version_ | 1797752140404359168 |
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| author | Elena Moreno-Giménez Elena Moreno-Giménez Mónica Gandía Zara Sáez Paloma Manzanares Lynne Yenush Diego Orzáez Jose F. Marcos Sandra Garrigues |
| author_facet | Elena Moreno-Giménez Elena Moreno-Giménez Mónica Gandía Zara Sáez Paloma Manzanares Lynne Yenush Diego Orzáez Jose F. Marcos Sandra Garrigues |
| author_sort | Elena Moreno-Giménez |
| collection | DOAJ |
| description | Fungal synthetic biology is a rapidly expanding field that aims to optimize the biotechnological exploitation of fungi through the generation of standard, ready-to-use genetic elements, and universal syntax and rules for contributory use by the fungal research community. Recently, an increasing number of synthetic biology toolkits have been developed and applied to filamentous fungi, which highlights the relevance of these organisms in the biotechnology field. The FungalBraid (FB) modular cloning platform enables interchangeability of DNA parts with the GoldenBraid (GB) platform, which is designed for plants, and other systems that are compatible with the standard Golden Gate cloning and syntax, and uses binary pCAMBIA-derived vectors to allow Agrobacterium tumefaciens-mediated transformation of a wide range of fungal species. In this study, we have expanded the original FB catalog by adding 27 new DNA parts that were functionally validated in vivo. Among these are the resistance selection markers for the antibiotics phleomycin and terbinafine, as well as the uridine-auxotrophic marker pyr4. We also used a normalized luciferase reporter system to validate several promoters, such as PpkiA, P7760, Pef1α, and PafpB constitutive promoters, and PglaA, PamyB, and PxlnA inducible promoters. Additionally, the recently developed dCas9-regulated GB_SynP synthetic promoter collection for orthogonal CRISPR activation (CRISPRa) in plants has been adapted in fungi through the FB system. In general, the expansion of the FB catalog is of great interest to the scientific community since it increases the number of possible modular and interchangeable DNA assemblies, exponentially increasing the possibilities of studying, developing, and exploiting filamentous fungi. |
| first_indexed | 2024-03-12T16:58:57Z |
| format | Article |
| id | doaj.art-9779e0e3b0e043538b47783861e845ce |
| institution | Directory Open Access Journal |
| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-03-12T16:58:57Z |
| publishDate | 2023-08-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-9779e0e3b0e043538b47783861e845ce2023-08-07T18:19:04ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852023-08-011110.3389/fbioe.2023.12228121222812FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungiElena Moreno-Giménez0Elena Moreno-Giménez1Mónica Gandía2Zara Sáez3Paloma Manzanares4Lynne Yenush5Diego Orzáez6Jose F. Marcos7Sandra Garrigues8Food Biotechnology Department, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Valencia, SpainInstituto de Biología Molecular y Celular de Plantas (IBMCP), Consejo Superior de Investigaciones Científicas (CSIC)-Universitat Politècnica de València (UPV), Valencia, SpainPreventive Medicine and Public Health, Food Science, Toxicology and Forensic Medicine Department. Faculty of Pharmacy. Universitat de València. Vicente Andrés Estellés s/n, Valencia, SpainFood Biotechnology Department, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Valencia, SpainFood Biotechnology Department, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Valencia, SpainInstituto de Biología Molecular y Celular de Plantas (IBMCP), Consejo Superior de Investigaciones Científicas (CSIC)-Universitat Politècnica de València (UPV), Valencia, SpainInstituto de Biología Molecular y Celular de Plantas (IBMCP), Consejo Superior de Investigaciones Científicas (CSIC)-Universitat Politècnica de València (UPV), Valencia, SpainFood Biotechnology Department, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Valencia, SpainFood Biotechnology Department, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Valencia, SpainFungal synthetic biology is a rapidly expanding field that aims to optimize the biotechnological exploitation of fungi through the generation of standard, ready-to-use genetic elements, and universal syntax and rules for contributory use by the fungal research community. Recently, an increasing number of synthetic biology toolkits have been developed and applied to filamentous fungi, which highlights the relevance of these organisms in the biotechnology field. The FungalBraid (FB) modular cloning platform enables interchangeability of DNA parts with the GoldenBraid (GB) platform, which is designed for plants, and other systems that are compatible with the standard Golden Gate cloning and syntax, and uses binary pCAMBIA-derived vectors to allow Agrobacterium tumefaciens-mediated transformation of a wide range of fungal species. In this study, we have expanded the original FB catalog by adding 27 new DNA parts that were functionally validated in vivo. Among these are the resistance selection markers for the antibiotics phleomycin and terbinafine, as well as the uridine-auxotrophic marker pyr4. We also used a normalized luciferase reporter system to validate several promoters, such as PpkiA, P7760, Pef1α, and PafpB constitutive promoters, and PglaA, PamyB, and PxlnA inducible promoters. Additionally, the recently developed dCas9-regulated GB_SynP synthetic promoter collection for orthogonal CRISPR activation (CRISPRa) in plants has been adapted in fungi through the FB system. In general, the expansion of the FB catalog is of great interest to the scientific community since it increases the number of possible modular and interchangeable DNA assemblies, exponentially increasing the possibilities of studying, developing, and exploiting filamentous fungi.https://www.frontiersin.org/articles/10.3389/fbioe.2023.1222812/fullfungal synthetic biologyGoldenBraidpromotersselection markersluciferase-based reporter systemCRISPR activation |
| spellingShingle | Elena Moreno-Giménez Elena Moreno-Giménez Mónica Gandía Zara Sáez Paloma Manzanares Lynne Yenush Diego Orzáez Jose F. Marcos Sandra Garrigues FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi Frontiers in Bioengineering and Biotechnology fungal synthetic biology GoldenBraid promoters selection markers luciferase-based reporter system CRISPR activation |
| title | FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi |
| title_full | FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi |
| title_fullStr | FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi |
| title_full_unstemmed | FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi |
| title_short | FungalBraid 2.0: expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi |
| title_sort | fungalbraid 2 0 expanding the synthetic biology toolbox for the biotechnological exploitation of filamentous fungi |
| topic | fungal synthetic biology GoldenBraid promoters selection markers luciferase-based reporter system CRISPR activation |
| url | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1222812/full |
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