Flexible gateway constructs for functional analyses of genes in plant pathogenic fungi

• Published in: Fungal genetics and biology Vol. 79; pp. 186 - 192
• Main Authors: Mehrabi, Rahim, Mirzadi Gohari, Amir, da Silva, Gilvan Ferreira, Steinberg, Gero, Kema, Gert H.J., de Wit, Pierre J.G.M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.06.2015
• Subjects: Binary vectors; Drug Resistance, Fungal; Fungi - genetics; Fungi - pathogenicity; Fungi - physiology; Gateway technology; Gene deletion; Gene Expression; Gene Targeting - methods; Genes, Fungal; Genetic Vectors; Genetics, Microbial - methods; GFP; Luminescent Proteins - analysis; Luminescent Proteins - genetics; Molecular Biology - methods; Plants - microbiology; RFP; Selection, Genetic; Staining and Labeling - methods; Transformation, Genetic; Triticum aestivum; Zymoseptoria tritici; GFP; Zymoseptoria tritici; Gene deletion; Binary vectors; RFP; Gateway technology
• ISSN: 1087-1845; 1096-0937
• DOI: 10.1016/j.fgb.2015.03.016

•The 22 entry vectors based on the Gateway vector methodology were developed.•Applications include up to triple gene deletion, overexpression, GFP or RFP labelling.•Vectors are a user-friendly tool for functional analyses of genes in fungi.•A number of these constructs was validated in Zymoseptoria tritici. Genetic manipulation of fungi requires quick, low-cost, efficient, high-throughput and molecular tools. In this paper, we report 22 entry constructs as new molecular tools based on the Gateway technology facilitating rapid construction of binary vectors that can be used for functional analysis of genes in fungi. The entry vectors for single, double or triple gene-deletion mutants were developed using hygromycin, geneticin and nourseothricin resistance genes as selection markers. Furthermore, entry vectors containing green fluorescent (GFP) or red fluorescent (RFP) in combination with hygromycin, geneticin or nourseothricin selection markers were generated. The latter vectors provide the possibility of gene deletion and simultaneous labelling of the fungal transformants with GFP or RFP reporter genes. The applicability of a number of entry vectors was validated in Zymoseptoria tritici, an important fungal wheat pathogen.