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Genetic transformation methods have recently been described; however, molecular tools for the functional study of genes have been scarcely reported for those fungi. In this work, we demonstrated that gene deletion and generation of the null mutant by homologous recombination are achievable for Fonsecaea pedrosoi by the use of two approaches: use of double-joint PCR for cassette construction, followed by delivery of the split-marker by biolistic transformation. Through in silico analyses, we identified that F. pedrosoi presents the complete enzymatic apparatus required for tryptophan (trp) biosynthesis. The gene encoding a tryptophan synthase trpB \u2014which converts chorismate to trp\u2014was disrupted. The \u0394trpB auxotrophic mutant can grow with external trp supply, but germination, viability of conidia, and radial growth are defective compared to the wild-type and reconstituted strains. The use of 5-FAA for selection of trp- phenotypes and for counter-selection of strains carrying the trp gene was also demonstrated. The molecular tools for the functional study of genes, allied to the genetic information from genomic databases, significantly boost our understanding of the biology and pathogenicity of CBM causative agents.<\/jats:p>","DOI":"10.3390\/jof9020224","type":"journal-article","created":{"date-parts":[[2023,2,9]],"date-time":"2023-02-09T01:37:07Z","timestamp":1675906627000},"page":"224","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Expanding the Toolbox for Functional Genomics in Fonsecaea pedrosoi: The Use of Split-Marker and Biolistic Transformation for Inactivation of Tryptophan Synthase (trpB) Gene"],"prefix":"10.3390","volume":"9","author":[{"given":"Lu\u00edsa Dan","family":"Favilla","sequence":"first","affiliation":[{"name":"Laboratory of Applied Immunology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Graduate Program in Molecular Biology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tatiana Sobianski","family":"Herman","sequence":"additional","affiliation":[{"name":"Laboratory of Applied Immunology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Graduate Program in Molecular Patology, Faculty of Medicine, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Camila da Silva","family":"Goersch","sequence":"additional","affiliation":[{"name":"Laboratory of Applied Immunology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Graduate Program in Microbial Biology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rosangela Vieira","family":"de Andrade","sequence":"additional","affiliation":[{"name":"Graduate Program of Genomic Sciences and Biotechnology, Catholic University of Brasilia, Campus Asa Norte, Asa Norte, Federal District, Taguatinga 70790-160, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria Sueli Soares","family":"Felipe","sequence":"additional","affiliation":[{"name":"Graduate Program of Genomic Sciences and Biotechnology, Catholic University of Brasilia, Campus Asa Norte, Asa Norte, Federal District, Taguatinga 70790-160, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3323-5300","authenticated-orcid":false,"given":"Anam\u00e9lia Lorenzetti","family":"Bocca","sequence":"additional","affiliation":[{"name":"Laboratory of Applied Immunology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Graduate Program in Molecular Biology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Graduate Program in Molecular Patology, Faculty of Medicine, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2814-7501","authenticated-orcid":false,"given":"Larissa","family":"Fernandes","sequence":"additional","affiliation":[{"name":"Laboratory of Applied Immunology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Graduate Program in Microbial Biology, Institute of Biology, Campus Darcy Ribeiro, University of Bras\u00edlia, Asa Norte, Federal District, Brasilia 70910-900, Brazil"},{"name":"Centro Metropolitano, Faculty of Ceil\u00e2ndia, Campus UnB Ceil\u00e2ndia, University of Bras\u00edlia, Ceil\u00e2ndia Sul, Federal District, Brasilia 72220-275, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,2,8]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"570","DOI":"10.1111\/j.1574-6976.2007.00077.x","article-title":"Biology and Pathogenesis of Fonsecaea pedrosoi, the Major Etiologic Agent of Chromoblastomycosis","volume":"31","author":"Santos","year":"2007","journal-title":"FEMS Microbiol. 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