A TRP1-marker-based system for gene complementation, overexpression, reporter gene expression and gene modification in Candida glabrata
| Autor: | Sascha Brunke, Bernhard Hube, Marcel Sprenger, Lydia Kasper |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
green fluorescent protein
Locus (genetics) Candida glabrata Applied Microbiology and Biotechnology Microbiology Genome gene complementation 03 medical and health sciences TRP1 Genes Reporter Candida albicans Gene 030304 developmental biology Genetics 0303 health sciences Reporter gene AcademicSubjects/SCI01150 biology 030306 microbiology Genetic Complementation Test Tryptophan ovalbumin General Medicine biology.organism_classification Corpus albicans Complementation urea amidolyase Mutation Genetic Engineering Research Article overexpression |
| Zdroj: | FEMS Yeast Research |
| ISSN: | 1567-1364 |
| Popis: | Although less prevalent than its relative Candida albicans, the yeast Candida glabrata is a successful pathogen of humans, which causes life-threatening candidiasis. It is thus vital to understand the pathogenicity mechanisms and contributing genes in C. glabrata. However, gene complementation as a tool for restoring the function of a previously deleted gene is not standardized in C. glabrata, and it is less frequently used than in C. albicans. In this study, we established a gene complementation strategy using genomic integration at the TRP1 locus. We prove that our approach can not only be used for integration of complementation cassettes, but also for overexpression of markers like fluorescent proteins and the antigen ovalbumin, or of potential pathogenicity-related factors like the biotin transporter gene VHT1. With urea amidolyase Dur1,2 as an example, we demonstrate the application of the gene complementation approach for the expression of sequence-modified genes. With this approach, we found that a lysine-to-arginine mutation in the biotinylation motif of Dur1,2 impairs urea-dependent growth of C. glabrata and C. albicans. Taken together, the TRP1-based gene complementation approach is a valuable tool for investigating novel gene functions and for elucidating their role in the pathobiology of C. glabrata. Introduction of a gene complementation and reporter gene expression strategy for the human-pathogenic yeast Candida glabrata, based on genomic integration at the TRP1 locus. |
| Databáze: | OpenAIRE |
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