Establishment of an efficient RNA silencing system in Trichoderma koningii using DsRed as a reporter
Language English Country United States Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Fluorescence MeSH
- Gene Knockdown Techniques methods MeSH
- Real-Time Polymerase Chain Reaction MeSH
- Luminescent Proteins analysis genetics MeSH
- RNA, Small Interfering genetics MeSH
- Genetics, Microbial methods MeSH
- Recombination, Genetic MeSH
- Genes, Reporter MeSH
- RNA Interference * MeSH
- Gene Expression Profiling MeSH
- Transformation, Genetic MeSH
- Trichoderma genetics MeSH
- Blotting, Western MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- fluorescent protein 583 MeSH Browser
- Luminescent Proteins MeSH
- RNA, Small Interfering MeSH
We aimed to establish an efficient RNA interference (RNAi) system in the industrially important filamentous fungus Trichoderma koningii using the DsRed protein as a reporter of the silencing process. To accomplish this, a DsRed expression cassette was transformed into T. koningii, and a recombinant strain that stably expressed DsRed was obtained. Next, a vector-directing expression of a DsRed hairpin RNA was constructed and transformed into the T. koningii recipient strain. Approximately 79 % of transformants displayed a decrease in DsRed fluorescence, and expression of DsRed in some transformants appeared to be fully suppressed. Characterization of randomly selected transformants by genomic DNA PCR analysis, real-time PCR quantification, and western blot confirmed downregulation of gene expression at different levels. The RNA silencing approach described here for T. koningii is effective, and the DsRed reporter gene provides a convenient tool for identification of silenced fungal transformants by their DsRed fluorescence compared to the control strain. The results of this study demonstrate the power of RNAi in T. koningii, which supports the use of this technology for strain development programs and functional genomics studies in industrial fungal strains.
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