Bidirectional-Genetics Platform, a Dual-Purpose Mutagenesis Strategy for Filamentous Fungi

Park, Jaejin; Lee, Yong-Hwan

doi:10.1128/ec.00234-13

Cited by 7 publications

(5 citation statements)

References 61 publications

(55 reference statements)

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“…Recycled ectopic transformants were generated in a BiG platform in M. oryzae KJ201 (Park and Lee, 2013). About 11% (128/1139) of mutants in this collection showed defects in vegetative growth, melanization and asexual reproduction (Park and Lee, 2013; Table 1).…”

Section: The Bidirectional Genetics (Big) Collectionmentioning

confidence: 99%

“…Recycled ectopic transformants were generated in a BiG platform in M. oryzae KJ201 (Park and Lee, ). About 11% (128/1139) of mutants in this collection showed defects in vegetative growth, melanization and asexual reproduction (Park and Lee, ; Table ). These BiG mutants were constructed by TGD‐based forward genetics, and reverse genetics involving the recycling of ectopic mutant isolates generated by random integration in MGG_09225 and MGG_14496 genes.…”

Section: Mutant Collectionsmentioning

confidence: 99%

“…The application of the BiG platform was verified by the assessment of disruptions in the putative histidinol‐phosphate aminotransferase gene, the promoter region of a homologue of S. cerevisiae vacuolar sorting‐associated protein 74 gene and a region homologous to the N. crassa frequency protein encoding gene. Park and Lee () provided a detailed technical evaluation of this collection for use in fungal genetic research. More mutant collections are yet to be constructed using this platform.…”

Section: Mutant Collectionsmentioning

confidence: 99%

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Large‐scale molecular genetic analysis in plant‐pathogenic fungi: a decade of genome‐wide functional analysis

Motaung

Saitoh

Tsilo

2017

Molecular Plant Pathology

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Plant-pathogenic fungi cause diseases to all major crop plants world-wide and threaten global food security. Underpinning fungal diseases are virulence genes facilitating plant host colonization that often marks pathogenesis and crop failures, as well as an increase in staple food prices. Fungal molecular genetics is therefore the cornerstone to the sustainable prevention of disease outbreaks. Pathogenicity studies using mutant collections provide immense function-based information regarding virulence genes of economically relevant fungi. These collections are rich in potential targets for existing and new biological control agents. They contribute to host resistance breeding against fungal pathogens and are instrumental in searching for novel resistance genes through the identification of fungal effectors. Therefore, functional analyses of mutant collections propel gene discovery and characterization, and may be incorporated into disease management strategies. In the light of these attributes, mutant collections enhance the development of practical solutions to confront modern agricultural constraints. Here, a critical review of mutant collections constructed by various laboratories during the past decade is provided. We used Magnaporthe oryzae and Fusarium graminearum studies to show how mutant screens contribute to bridge existing knowledge gaps in pathogenicity and fungal-host interactions.

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Section: The Bidirectional Genetics (Big) Collectionmentioning

confidence: 99%

Section: Mutant Collectionsmentioning

confidence: 99%

Section: Mutant Collectionsmentioning

confidence: 99%

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Large‐scale molecular genetic analysis in plant‐pathogenic fungi: a decade of genome‐wide functional analysis

Motaung

Saitoh

Tsilo

2017

Molecular Plant Pathology

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“…MoFRQ was previously identified by a bidirectional genetic platform. An insertional mutant of MoFRQ displayed no apparent phenotypes except abnormal conidiophores (Park and Lee, ). In contrast, the deletion of MoFRQ caused slow growth, reduced conidiation, lower conidial germination and appressorial formation and reduced virulence in our work, indicating that MoFRQ is involved in development and differentiation in M. oryzae .…”

Section: Discussionmentioning

confidence: 99%

“…We deleted a genomic region, which was previously annotated as MGG_00903. Differently, the insertional mutant of MoFRQ had mutation in the C‐terminal region only (Park and Lee, ).…”

Section: Discussionmentioning

confidence: 99%

F‐box proteins MoFwd1, MoCdc4 and MoFbx15 regulate development and pathogenicity in the rice blast fungus Magnaporthe oryzae

Shi

Chen

Zhu

et al. 2019

Environmental Microbiology

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Summary The Skp1‐Cul1‐F‐box‐protein (SCF) ubiquitin ligases are important parts of the ubiquitin system controlling many cellular biological processes in eukaryotes. However, the roles of SCF ubiquitin ligases remain unclear in phytopathogenic Magnaporthe oryzae. Here, we cloned 24 F‐box proteins and confirmed that 17 proteins could interact with MoSkp1, showing their potential to participate in SCF complexes. To determine their functions, null mutants of 21 F‐box‐containing genes were created. Among them, the F‐box proteins MoFwd1, MoCdc4 and MoFbx15 were found to be required for growth, development and full virulence. Fluorescent‐microscopy observations demonstrated that both MoFbx15 and MoCdc4 were localized to the nucleus, compared with MoFwd1, which was distributed in the cytosol. MoCdc4 and MoFwd1 bound to MoSkp1 via the F‐box domain, the deletion of which abrogated their function. Race tube and qRT‐PCR assays confirmed that MoFwd1 was involved in circadian rhythm by regulating transcription and protein stability of the core circadian clock regulator MoFRQ. Moreover, MoFWD1 also orchestrates conidial germination by influencing conidial amino acids pools and oxidative stress release. Overall, our results indicate that SCF ubiquitin ligases play indispensable roles in development and pathogenicity in M. oryzae.

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Restriction enzyme-mediated insertional mutagenesis: an efficient method of Rosellinia necatrix transformation

2017

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Rosellinia necatrix: causing root rot disease is a very destructive pathogen of woody plants and is responsible for yield losses to a large number of fruit trees. The genetic analysis of this pathogen has not been picked up because of difficulty in generating mutations in Rosellinia necatrix for many reasons. A number of methods have been proposed for inducing mutations in Rosellinia necatrix but none of them proved worth because of very low transformation efficiencies. Here, we propose an efficient method for Rosellinia necatrix protoplast production, where protoplasts in the tune of 10 per ml can be easily generated. We also propose a restriction enzyme-mediated integration (REMI)-based methods for efficient transformation of Rosellinia necatrix. In the present study, an approximate of 800 transformants was obtained from 5 μg of linearized plasmid. Out of 47 single spored transformants analyzed, only 33 showed hygromycin gene amplification using PCR and only 19 transformants showed single gene integration in southern hybridization, which accounted for single gene integration percentage of 42%, highest amongst all the previous reports on Rosellinia necatrix transformations. Some of the transformants studied for pathogenicity phenotype also showed a marked reduction in pathogenicity. Thus, in the present investigation, 42% single gene integrations among the transformed colonies can be considered as excellent transformation efficiency.

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Bidirectional-Genetics Platform, a Dual-Purpose Mutagenesis Strategy for Filamentous Fungi

Cited by 7 publications

References 61 publications

Large‐scale molecular genetic analysis in plant‐pathogenic fungi: a decade of genome‐wide functional analysis

Large‐scale molecular genetic analysis in plant‐pathogenic fungi: a decade of genome‐wide functional analysis

F‐box proteins MoFwd1, MoCdc4 and MoFbx15 regulate development and pathogenicity in the rice blast fungus Magnaporthe oryzae

Restriction enzyme-mediated insertional mutagenesis: an efficient method of Rosellinia necatrix transformation

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