Phenotypic Analysis of Neurospora crassa Gene Deletion Strains

2011 ◽  
pp. 191-198 ◽  
Author(s):  
Gloria E. Turner
Keyword(s):  
Neurospora Crassa ◽  
Gene Deletion ◽  
Phenotypic Analysis

scholarly journals High-Throughput Construction of Gene Deletion Cassettes for Generation of Neurospora crassa Knockout Strains

2010 ◽  
pp. 33-40 ◽  
Author(s):  
Patrick D. Collopy ◽  
Hildur V. Colot ◽  
Gyungsoon Park ◽  
Carol Ringelberg ◽  
Christopher M. Crew ◽  
...  
Keyword(s):  
Neurospora Crassa ◽  
High Throughput ◽  
Gene Deletion

Direct cellobiose production from cellulose using sextuple beta-glucosidase gene deletion Neurospora crassa mutants

2013 ◽  
Vol 52 (3) ◽  
pp. 184-189 ◽  
Author(s):  
Weihua Wu ◽  
Amanda Hildebrand ◽  
Takao Kasuga ◽  
Xiaochao Xiong ◽  
Zhiliang Fan
Keyword(s):  
Neurospora Crassa ◽  
Gene Deletion ◽  
Beta Glucosidase

Comprehensive phenotypic analysis of single-gene deletion and overexpression strains of Saccharomyces cerevisiae

Yeast ◽  
2011 ◽  
Vol 28 (5) ◽  
pp. 349-361 ◽  
Author(s):  
Katsunori Yoshikawa ◽  
Tadamasa Tanaka ◽  
Yoshihiro Ida ◽  
Chikara Furusawa ◽  
Takashi Hirasawa ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Deletion ◽  
Single Gene ◽  
Phenotypic Analysis ◽  
Single Gene Deletion

scholarly journals The Predicted G-Protein-Coupled Receptor GPR-1 Is Required for Female Sexual Development in the Multicellular Fungus Neurospora crassa

2006 ◽  
Vol 5 (9) ◽  
pp. 1503-1516 ◽  
Author(s):  
Svetlana Krystofova ◽  
Katherine A. Borkovich
Keyword(s):  
Neurospora Crassa ◽  
G Protein ◽  
Sexual Development ◽  
Fluorescent Protein ◽  
Protein Fusion ◽  
Phenotypic Analysis ◽  
Reproductive Structures ◽  
Epistasis Analysis ◽  
Green Fluorescent ◽  
G Protein Coupled

ABSTRACTG-protein-coupled receptors (GPCRs) control important aspects of asexual and sexual development in eukaryotic organisms. We have identified a predicted GPCR in the filamentous fungusNeurospora crassawith similarity to cyclic AMP-receptor like GPCRs fromDictyostelium discoideumand GCR1 fromArabidopsis thaliana. Expression ofgpr-1is highest in female reproductive structures, and deletion ofgpr-1leads to defects during sexual development. Unfertilized female structures (protoperithecia) fromΔgpr-1strains are weakly pigmented, small, and submerged in the agar. The perithecia produced after fertilization have deformed beaks that lack ostioles, the openings through which ascospores are discharged. Localization studies using a GPR-1-green fluorescent protein fusion protein showed that GPR-1 is targeted to female reproductive structures. Genetic epistasis experiments with the three Gα genes were inconclusive due to the early block in mating exhibited by Δgna-1strains. Phenotypic analysis of mutants from a high-throughputN. crassaknockout project allowed identification of BEK-1, a homeodomain transcription factor that is a potential target of GPR-1. The perithecial defects ofΔbek-1strains are similar to those of theΔgpr-1strain, and epistasis analysis indicates thatbek-1could function downstream ofgpr-1during postfertilization events. The effect must be posttranscriptional, asbek-1transcript levels are not affected inΔgpr-1strains. The lack of ostioles inΔgpr-1and Δbek-1mutants has an undesirable effect on the ability to spread progeny (ascospores) by the normal ejection mechanism and would severely compromise the fitness of these strains in nature.

scholarly journals Characterization of Single Gene Deletion Mutants Affecting Alternative Oxidase Production in Neurospora crassa: Role of the yvh1 Gene

2020 ◽  
Vol 8 (8) ◽  
pp. 1186
Author(s):  
Adrien Beau Desaulniers ◽  
Nishka Kishore ◽  
Kelly Adames ◽  
Frank E. Nargang
Keyword(s):  
Neurospora Crassa ◽  
Gene Deletion ◽  
Alternative Oxidase ◽  
Single Gene ◽  
Ribosome Assembly ◽  
Mitochondrial Translation ◽  
Multiple Phenotypes ◽  
Transport Chain ◽  
Zinc Binding Domain ◽  
Single Gene Deletion

The Neurospora crassa AOD1 protein is a mitochondrial alternative oxidase that passes electrons directly from ubiquinol to oxygen. The enzyme is encoded by the nuclear aod-1 gene and is produced when the standard electron transport chain is inhibited. We previously identified eleven strains in the N. crassa single gene deletion library that were severely deficient in their ability to produce AOD1 when grown in the presence of chloramphenicol, an inhibitor of mitochondrial translation that is known to induce the enzyme. Three mutants affected previously characterized genes. In this report we examined the remaining mutants and found that the deficiency of AOD1 was due to secondary mutations in all but two of the strains. One of the authentic mutants contained a deletion of the yvh1 gene and was found to have a deficiency of aod-1 transcripts. The YVH1 protein localized to the nucleus and a post mitochondrial pellet from the cytoplasm. A zinc binding domain in the protein was required for rescue of the AOD1 deficiency. In other organisms YVH1 is required for ribosome assembly and mutants have multiple phenotypes. Lack of YVH1 in N. crassa likely also affects ribosome assembly leading to phenotypes that include altered regulation of AOD1 production.

scholarly journals Towards a Systems Approach in the Genetic Analysis of Archaea: Accelerating Mutant Construction and Phenotypic Analysis inHaloferax volcanii

Archaea ◽  
2010 ◽  
Vol 2010 ◽  
pp. 1-11 ◽  
Author(s):  
Ian K. Blaby ◽  
Gabriela Phillips ◽  
Crysten E. Blaby-Haas ◽  
Kevin S. Gulig ◽  
Basma El Yacoubi ◽  
...  
Keyword(s):  
High Throughput ◽  
Gene Deletion ◽  
Systems Approach ◽  
Essential Gene ◽  
Gene Knockout ◽  
Deletion Mutants ◽  
Mutant Library ◽  
Phenotypic Analysis ◽  
Genome Wide ◽  
A Genome

With the availability of a genome sequence and increasingly sophisticated genetic tools,Haloferax volcaniiis becoming a model for both Archaea and halophiles. In order forH. volcaniito reach a status equivalent toEscherichia coli, Bacillus subtilis, orSaccharomyces cerevisiae, a gene knockout collection needs to be constructed in order to identify the archaeal essential gene set and enable systematic phenotype screens. A streamlined gene-deletion protocol adapted for potential automation was implemented and used to generate 22H. volcaniideletion strains and identify several potentially essential genes. These gene deletion mutants, generated in this and previous studies, were then analyzed in a high-throughput fashion to measure growth rates in different media and temperature conditions. We conclude that these high-throughput methods are suitable for a rapid investigation of anH. volcaniimutant library and suggest that they should form the basis of a larger genome-wide experiment.

scholarly journals Overlapping Functions for Two G Protein α Subunits in Neurospora crassa

Genetics ◽  
1997 ◽  
Vol 147 (1) ◽  
pp. 137-145 ◽  
Author(s):  
Rudeina A Baasiri ◽  
Xiaohui Lu ◽  
Patricia S Rowley ◽  
Gloria E Turner ◽  
Katherine A Borkovich
Keyword(s):  
Neurospora Crassa ◽  
G Protein ◽  
Sexual Cycle ◽  
Fusion Activity ◽  
Heterotrimeric G Proteins ◽  
Phenotypic Analysis ◽  
Activating Mutations ◽  
Eukaryotic Microbes ◽  
Α Subunits ◽  
G Protein Α Subunits

Abstract Heterotrimeric G proteins, consisting of α, β and γ subunits, mediate a variety of signaling pathways in eukaryotes. We have previously identified two genes, gna-1 and gna-2, that encode G protein α subunits in the filamentous fungus Neurospora crassa. Mutation of gna-1 results in female infertility and sensitivity to hyperosmotic media. In this study, we investigate the expression and functions of gna-2. Results from Western analysis and measurements of gna-2 promoter-lacZ fusion activity indicate that gna-2 is expressed during the vegetative and sexual cycle of N. crassa in both A and a mating types. Activating mutations predicted to abolish the GTPase activity of GNA-2 cause subtle defects in aerial hyphae formation and conidial germination. Extensive phenotypic analysis of Δgna-2 strains did not reveal abnormalities during vegetative or sexual development. In contrast, deletion of gna-2 in a Δgna-1 strain accentuates the Δgna-1 phenotypes. Δgna-1 Δgna-2 strains have a slower rate of hyphal apical extension than Δgna-1 strains on hyperosmotic media. Moreover, Δgna-1 Δgna-2 mutants have more pronounced defects in female fertility than Δgna-1 strains. We propose that gna-1 and gna-2 have overlapping functions and may constitute a gene family. This is the first report of G protein α subunits with overlapping functions in eukaryotic microbes.

scholarly journals A Gateway platform for functional genomics inHaloferax volcanii: deletion of three tRNA modification genes

Archaea ◽  
2009 ◽  
Vol 2 (4) ◽  
pp. 211-219 ◽  
Author(s):  
Basma El Yacoubi ◽  
Gabriela Phillips ◽  
Ian K. Blaby ◽  
Crysten E. Haas ◽  
Yulien Cruz ◽  
...  
Keyword(s):  
Gene Deletion ◽  
Genetic Manipulation ◽  
Model Organism ◽  
Rna Modification ◽  
Deletion Mutants ◽  
Trna Modification ◽  
Gtp Cyclohydrolase I ◽  
Phenotypic Analysis ◽  
Targeted Gene Deletion ◽  
Folate Biosynthesis

In part due to the existence of simple methods for its cultivation and genetic manipulation,Haloferax volcaniiis a major archaeal model organism. It is the only archaeon for which the whole set of post-transcriptionally modified tRNAs has been sequenced, allowing for anin silicoprediction of all RNA modification genes present in the organism. One approach to check these predictions experimentally is via the construction of targeted gene deletion mutants. Toward this goal, an integrative “Gateway vector” that allows gene deletion inH. volcaniiuracil auxotrophs was constructed. The vector was used to delete three predicted tRNA modification genes: HVO_2001 (encoding an archaeal transglycosyl tranferase or arcTGT), which is involved in archeosine biosynthesis; HVO_2348 (encoding a newly discovered GTP cyclohydrolase I), which catalyzes the first step common to archaeosine and folate biosynthesis; and HVO_2736 (encoding a member of the COG1444 family), which is involved inN4-acetylcytidine (ac4C) formation. Preliminary phenotypic analysis of the deletion mutants was conducted, and confirmed all three predictions.

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