A high-throughput gene knockout procedure for Neurospora reveals functions for multiple transcription factors.
Hildur V. Colot,Gyungsoon Park,Gloria E. Turner,Carol S. Ringelberg,Christopher M. Crew,Liubov Litvinkova,Richard L. Weiss,Katherine A. Borkovich,Jay C. Dunlap +8 more
TL;DR: This study describes a method for rapidly creating knockout mutants in which it makes use of yeast recombinational cloning, Neurospora mutant strains deficient in nonhomologous end-joining DNA repair, custom-written software tools, and robotics.
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Abstract: The low rate of homologous recombination exhibited by wild-type strains of filamentous fungi has hindered development of high-throughput gene knockout procedures for this group of organisms. In this study, we describe a method for rapidly creating knockout mutants in which we make use of yeast recombinational cloning, Neurospora mutant strains deficient in nonhomologous end-joining DNA repair, custom-written software tools, and robotics. To illustrate our approach, we have created strains bearing deletions of 103 Neurospora genes encoding transcription factors. Characterization of strains during growth and both asexual and sexual development revealed phenotypes for 43% of the deletion mutants, with more than half of these strains possessing multiple defects. Overall, the methodology, which achieves high-throughput gene disruption at an efficiency >90% in this filamentous fungus, promises to be applicable to other eukaryotic organisms that have a low frequency of homologous recombination.
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Citations
Epigenetic Control of Phenotypic Plasticity in the Filamentous Fungus Neurospora crassa
TL;DR: It is shown that epigenetic mechanisms play a role in homeostasis and phenotypic plasticity of the fungus across a range of controlled environments, indicating that epigenetics regulation is context dependent and is not governed by general plasticity genes.
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A Homeobox Transcription Factor UvHOX2 Regulates Chlamydospore Formation, Conidiogenesis, and Pathogenicity in Ustilaginoidea virens.
Junjie Yu,Mina Yu,Tianqiao Song,Huijuan Cao,Xiayan Pan,Mingli Yong,Zhongqiang Qi,Yan Du,Rongsheng Zhang,Xiaole Yin,Yongfeng Liu +10 more
TL;DR: It is confirmed that UvHOX2 is located in the nuclei of U. virens, and the expression of UvHox2 was the strongest during the early stage of chlamydospore and conidium formation, and global transcription pattern was determined by RNA-seq.
The polyketide synthase gene pks4 is essential for sexual development and regulates fruiting body morphology in Sordaria macrospora
TL;DR: The data show that the polyketide synthase gene pks4 is essential for sexual development and plays a role in regulating fruiting body morphology, and correct expression levels of pKS4 are essential for wild type-like perithecia formation.
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Identification of PEX33, a novel component of the peroxisomal docking complex in the filamentous fungus Neurospora crassa.
David Managadze,Christian Würtz,Sebastian Wiese,Sebastian Wiese,Michael Schneider,Wolfgang Girzalsky,Helmut E. Meyer,Ralf Erdmann,Bettina Warscheid,Bettina Warscheid,Hanspeter Rottensteiner +10 more
TL;DR: The isolation of PEX14 complexes of Neurospora crassa revealed a defect of this strain in the biogenesis of glyoxysomes and Woronin bodies and a genuine novel peroxin.
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Regulation of Cell-to-Cell Communication and Cell Wall Integrity by a Network of MAP Kinase Pathways and Transcription Factors in Neurospora crassa
TL;DR: The data demonstrate that the MAK-1/ADV-1 andMAK-2/PP-1 pathways form a tight regulatory network that maintains cell integrity and mediates communication and cell fusion in N. crassa.
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TL;DR: A total of 6925 Saccharomyces cerevisiae strains were constructed, by a high-throughput strategy, each with a precise deletion of one of 2026 ORFs (more than one-third of the ORFs in the genome), finding that 17 percent were essential for viability in rich medium.
Molecular Bases for Circadian Clocks
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The genome sequence of the filamentous fungus Neurospora crassa
James E. Galagan,Sarah E. Calvo,Katherine A. Borkovich,Eric U. Selker,Nick O. Read,David B. Jaffe,William Fitzhugh,Li-Jun Ma,Serge Smirnov,Seth Purcell,Bushra Rehman,Timothy Elkins,Reinhard Engels,Shunguang Wang,Cydney B. Nielsen,Jonathan Butler,Matthew G. Endrizzi,Dayong Qui,Peter Ianakiev,Deborah Bell-Pedersen,Mary Anne Nelson,Margaret Werner-Washburne,Claude P. Selitrennikoff,John A. Kinsey,Edward L. Braun,Alex Zelter,Alex Zelter,Ulrich Schulte,Gregory O. Kothe,Gregory Jedd,Werner Mewes,Chuck Staben,Edward M. Marcotte,David Greenberg,Alice Roy,Karen Foley,Jerome Naylor,Nicole Stange-Thomann,Robert Barrett,Sante Gnerre,Michael Kamal,Manolis Kamvysselis,Evan Mauceli,Cord Bielke,Stephen Rudd,Dmitrij Frishman,Svetlana Krystofova,Carolyn G. Rasmussen,Robert L. Metzenberg,David D. Perkins,Scott Kroken,Carlo Cogoni,Giuseppe Macino,David E. A. Catcheside,Weixi Li,Robert J. Pratt,Stephen A. Osmani,Colin P.C. DeSouza,Louise Glass,Marc J. Orbach,J. Andrew Berglund,Rodger B. Voelker,Oded Yarden,Michael Plamann,Stephan Seiler,Jay C. Dunlap,Alan Radford,Rodolfo Aramayo,Donald O. Natvig,Lisa A. Alex,Gertrud Mannhaupt,Daniel J. Ebbole,Michael Freitag,Ian T. Paulsen,Matthew S. Sachs,Eric S. Lander,Chad Nusbaum,Bruce W. Birren +77 more
TL;DR: A high-quality draft sequence of the N. crassa genome is reported, suggesting that RIP has had a profound impact on genome evolution, greatly slowing the creation of new genes through genomic duplication and resulting in a genome with an unusually low proportion of closely related genes.
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