title: A modular plasmid system for protein co-localization and bimolecular fluorescence complementation in filamentous fungi.
abstract: To elucidate the function of a protein, it is crucial to know its subcellular location and its interaction partners. Common approaches to resolve those questions rely on the genetic tagging of the gene-of-interest (GOI) with fluorescent reporters. To determine the location of a tagged protein, it may be co-localized with tagged marker proteins. The interaction of two proteins under investigation is often analysed by tagging both with the C- and N-terminal halves of a fluorescent protein. In fungi, the tagged GOI are commonly introduced by serial transformation with plasmids harbouring a single tagged GOI and subsequent selection of suitable strains. In this study, a plasmid system is presented that allows the tagging of several GOI on a single plasmid. This novel double tagging plasmid system (DTPS) allows a much faster and less laborious generation of double-labelled fungal strains when compared with conventional approaches. The DTPS also enables the combination of as many tagged GOI as desired and a simple exchange of existing tags. Furthermore, new tags can be introduced smoothly into the system. In conclusion, the DTPS allows an efficient tagging of GOI with a high degree of flexibility and therefore accelerates functional analysis of proteins in vivo.
authors.affiliation: Plant Nutrition Laboratory, Faculty of Natural Sciences III, Institute of Agricultural and Nutritional Sciences, Martin Luther University of Halle-Wittenberg, 06099, Halle (Saale), Germany.
authors.forename: Mario
authors.initials: M
authors.lastname: Lange
authors.forename: Ely
authors.initials: E
authors.lastname: Oliveira-Garcia
authors.forename: Holger B
authors.initials: HB
authors.lastname: Deising
authors.forename: Edgar
authors.initials: E
authors.lastname: Peiter
mesh_terms: D020231:Colletotrichum / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D005453:Fluorescence; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D018390:Gene Targeting; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D021381:Protein Transport; D011993:Recombinant Fusion Proteins; D000097573:Red Fluorescent Protein
chemical_list: D005656:Fungal Proteins; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D000097573:Red Fluorescent Protein