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Bidirectional-Genetics Platform, a Dual-Purpose Mutagenesis Strategy for Filamentous Fungi
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 Authours
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Jaejin Park, Yong-Hwan Lee |
| Title |
Bidirectional-Genetics Platform, a Dual-Purpose Mutagenesis Strategy for Filamentous Fungi |
| Journal |
Eukaryotic Cell, 2013 (12) ~ |
Abstract
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Rapidly increasing fungal genome sequences call for efficient ways of generating mutants to translate quickly gene sequences into their functions. A reverse genetic strategy via targeted gene replacement (TGR) has been inefficient for many filamentous fungi due to dominant production of undesirable ectopic transformants. Although large-scale random insertional mutagenesis via transformation (i.e., forward genetics) facilitates high-throughput uncovering of novel genes of interest, generating a huge number of transformants, which is necessary to ensure the likelihood of mutagenizing most genes, is time-consuming. We propose a new strategy, entitled the Bidirectional-Genetics (BiG) platform, which combines both forward and reverse genetic strategies by recycling ectopic transformants derived from TGR as a source for random insertional mutants. The BiG platform was evaluated using the rice blast fungus Magnaporthe oryzae as a model. Over 10% of >1,000 M. oryzae ectopic transformants, generated during disruption of specific genes, displayed abnormality in vegetative growth, pigmentation, and/or asexual reproduction. In this pool of putative mutants, we isolated insertional mutants with mutations in three genes involved in histidine biosynthesis (MoHIS5), vegetative growth (MoVPS74), or conidiophore formation (MoFRQ) (where ˇ°Moˇ± indicates ˇ°M. oryzaeˇ±), supporting the utility of this platform for systematic gene function studies. |
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