scholarly journals Marked Neurospora crassa strains for competition experiments and Bayesian methods for fitness estimates

2019 ◽  
Author(s):  
Ilkka Kronholm ◽  
Tereza Ormsby ◽  
Kevin J. McNaught ◽  
Eric U. Selker ◽  
Tarmo Ketola
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Relative Fitness ◽  
Fitness Effect ◽  
Type Locus ◽  
Competitive Fitness ◽  
Measured Strain ◽  
Different Strains ◽  
Suitable Marker ◽  
General Tool

AbstractThe filamentous fungus Neurospora crassa, a model microbial eukaryote, has a life cycle with many features that make it suitable for studying experimental evolution. However, it has lacked a general tool for estimating relative fitness of different strains in competition experiments. To remedy this need, we constructed N. crassa strains that contain a modified csr-1 locus and developed an assay for detecting the proportion of the marked strain using a post PCR high resolution melting assay. DNA extraction from spore samples can be performed on 96-well plates, followed by a PCR step, which allows many samples to be processed with ease. Furthermore, we suggest a Bayesian approach for estimating relative fitness from competition experiments that takes into account the uncertainty in measured strain proportions. We show that there is a fitness effect of the mating type locus, as mating type mat a has a higher competitive fitness than mat A. The csr-1* marker also has a small fitness effect, but is still a suitable marker for competition experiments. As a proof of concept, we estimate the fitness effect of the qde-2 mutation, a gene in the RNA interference pathway, and show that its competitive fitness is lower than what would be expected from its mycelial growth rate alone.

scholarly journals Marked Neurospora crassa Strains for Competition Experiments and Bayesian Methods for Fitness Estimates

2020 ◽  
Vol 10 (4) ◽  
pp. 1261-1270 ◽  
Author(s):  
Ilkka Kronholm ◽  
Tereza Ormsby ◽  
Kevin J. McNaught ◽  
Eric U. Selker ◽  
Tarmo Ketola
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Relative Fitness ◽  
Fitness Effect ◽  
Type Locus ◽  
Competitive Fitness ◽  
Measured Strain ◽  
Different Strains ◽  
Suitable Marker ◽  
General Tool

The filamentous fungus Neurospora crassa, a model microbial eukaryote, has a life cycle with many features that make it suitable for studying experimental evolution. However, it has lacked a general tool for estimating relative fitness of different strains in competition experiments. To remedy this need, we constructed N. crassa strains that contain a modified csr-1 locus and developed an assay for detecting the proportion of the marked strain using a post PCR high resolution melting assay. DNA extraction from spore samples can be performed on 96-well plates, followed by a PCR step, which allows many samples to be processed with ease. Furthermore, we suggest a Bayesian approach for estimating relative fitness from competition experiments that takes into account the uncertainty in measured strain proportions. We show that there is a fitness effect of the mating type locus, as mating type mat a has a higher competitive fitness than mat A. The csr-1* marker also has a small fitness effect, but is still a suitable marker for competition experiments. As a proof of concept, we estimate the fitness effect of the qde-2 mutation, a gene in the RNA interference pathway, and show that its competitive fitness is lower than what would be expected from its mycelial growth rate alone.

A SEARCH FOR COMPLEXITY AT THE MATING-TYPE LOCUS OF NEUROSPORA CRASSA

1973 ◽  
Vol 15 (3) ◽  
pp. 577-585 ◽  
Author(s):  
Dorothy Newmeyer ◽  
H. Branch Howe Jr. ◽  
Donna R. Galeazzi
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Adjacent Gene ◽  
Opposite Mating Type ◽  
Type Locus ◽  
Linked Gene ◽  
Heterokaryon Incompatibility ◽  
Mating Type Locus ◽  
Incompatibility Reaction ◽  
The Right

Evidence for complexity at the mating-type locus of Neurospora crassa was sought by selecting recombinants between closely linked markers on either side. All recombinants were tested for crossing ability, to test the hypothesis that the two mating-type alleles are actually closely linked self-sterile mutants; such tests should also detect subunits analogous to the α and β subunits of the A factor of Schizophyllum or Coprinus. No change in crossing ability was found among the 5,019 recombinants tested, representing 235,000 viable ascospores. The results indicate that if subunits exist, they are not more than 0.002 units apart. Twelve hundred and forty of the recombinants were tested in a way that should also have detected subunits analogous to the A and B factors of Schizophyllum and Coprinus, except that A and B would be closely linked. No such subunits were detected.N. crassa strains of opposite mating type are heterokaryon-incompatible during vegetative growth, and observations of various investigators have suggested that the heterokaryon incompatibility might be controlled by a separate closely-linked gene rather than by mating type itself. A sample of the recombinants was therefore tested for separation of the heterokaryon-incompatibility and crossing-compatibility functions. (Heterokaryon-incompatibility was scored by the presence of an incompatibility reaction in duplications heterozygous for mating type; this technique is simple and eliminates complications due to unlinked heterokaryon-incompatibility loci, several of which are known in N. crassa.) No separation was found. The results indicate that if an adjacent gene is responsible for the heterokaryon-incompatibility, it is not more than 0.0078 units from mating type, if on the left, and not more than 0.018 units from mating type, if on the right.

scholarly journals A Second Gene Controlling Allelic Recombination in Neurospora Crassa

1966 ◽  
Vol 19 (6) ◽  
pp. 1039 ◽  
Author(s):  
DG Catoheside
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Type Locus ◽  
Mating Type Locus

Experiments to determine whether rec-l, which increases allelic recombinationat the his-l locus in Neurospora crassa, also affects the am locus disclosed anothergene, rec-3. It appears that ree-l is specific to his-l and that ree-3 is specific to am, inthe sense that his-l is insensitive to rec-3 and am is insensitive to ree-l. The locus of rec-l is 18�9 units from the am his-l region; rec-3 is linked either to the am his-l region or to the mating type locus and 12�1 units from the relevant region.

Characterization of mat A-2, mat A-3 and ΔmatA Mating-Type Mutants of Neurospora crassa

Genetics ◽  
1998 ◽  
Vol 148 (3) ◽  
pp. 1069-1079 ◽  
Author(s):  
Adlane V-B Ferreira ◽  
Zhiqiang An ◽  
Robert L Metzenberg ◽  
N Louise Glass
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Sexual Development ◽  
Vegetative Growth ◽  
Double Mutant ◽  
Sexual Cycle ◽  
Wild Type ◽  
Type Locus ◽  
Isolation And Characterization

AbstractThe mating-type locus of Neurospora crassa regulates mating identity and entry into the sexual cycle. The mat A idiomorph encodes three genes, mat A-1, mat A-2, and mat A-3. Mutations in mat A-1 result in strains that have lost mating identity and vegetative incompatibility with mat a strains. A strain containing mutations in both mat A-2 and mat A-3 is able to mate, but forms few ascospores. In this study, we describe the isolation and characterization of a mutant deleted for mat (ΔmatA), as well as mutants in either mat A-2 or mat A-3. The ΔmatA strain is morphologically wild type during vegetative growth, but it is sterile and heterokaryon compatible with both mat A and mat a strains. The mat A-2 and mat A-3 mutants are also normal during vegetative growth, mate as a mat A strain, and produce abundant biparental asci in crosses with mat a, and are thus indistinguishable from a wild-type mat A strain. These data and the fact that the mat A-2 mat A-3 double mutant makes few asci with ascospores indicate that MAT A-2 and MAT A-3 are redundant and may function in the same pathway. Analysis of the expression of two genes (sdv-1 and sdv-4) in the various mat mutants suggests that the mat A polypeptides function in concert to regulate the expression of some sexual development genes.

scholarly journals Directed replacement of mt A by mt a-1 effects a mating type switch in Neurospora crassa.

Genetics ◽  
1994 ◽  
Vol 138 (1) ◽  
pp. 75-81 ◽  
Author(s):  
S Chang ◽  
C Staben
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Single Copy ◽  
Gene Replacement ◽  
Type Locus ◽  
Sexual Activities ◽  
Sexual Process ◽  
Type Switch ◽  
Type Gene ◽  
Necessary And Sufficient

Abstract To test the functions of a mating type genes, we developed an efficient strategy to select transformants of Neurospora crassa in which resident A mating type DNA was replaced by cloned DNA from the mt a idiomorph. Cloned a idiomorphic DNA could specify all functions, including fertility, of a mating type, but only when it replaced A DNA at the mating type locus. Only the mt a-1 region of the a idiomorph was necessary in order to specify a mating type. Gene replacement events involved the homologous sequences flanking the unique mating type idiomorphic DNA, resulting in apparently isogenic a and A strains. These isogenic strains were fertile when crossed with one another, indicating that no determinants outside the transforming DNA are necessary for fertility as a and that no host sequences of A strains interfere with fertility as a. One a replacement strain bore a duplication of the transforming mt a-1 and hph DNA. The duplication strain had unexpected properties. Although mating type segregated 1:1 in crosses of this strain to A, the duplicated regions were efficiently altered during the sexual process to generate a single copy in the progeny. No progeny were recovered that had undergone RIP (repeat induced point mutation) sufficient to inactivate the mt a-1 gene. We infer that the mt a-1 gene is necessary and sufficient to specify a mating type identity in all vegetative and sexual activities. Mt a-1 may also play an essential role in ascosporogenesis after fertilization.

scholarly journals GENES INFLUENCING SELECTIVE FERTILIZATION IN NEUROSPORA CRASSA

Genetics ◽  
1972 ◽  
Vol 70 (4) ◽  
pp. 511-519
Author(s):  
Takeshi Egashira ◽  
Kuzuo Nakamura
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Genetic Similarity ◽  
Mutant Gene ◽  
Opposite Mating Type ◽  
Cytoplasmic Effect ◽  
Mutual Attraction ◽  
Color Mutant ◽  
Different Strains

ABSTRACT The mutual attraction of conidia to protoperithecia of the opposite mating type was studied genetically in crosses where a mixture of conidia from two different strains, one of which was marked by an ascospore color mutant gene tan spore (ts), was applied to protoperithecia. Selective fertilization was measured as the frequency of perithecia fertilized by conidia from one strain in competition with conidia from another strain. Selective fertilization by a given strain varied throughout the range from 10 to 97% according to the strains of protoperithecial parent. The selective fertilization was revealed to be under the control of two or more loci, which appeared to have multiplicative action. No indication of a cytoplasmic effect on selective fertilization was obtained. The strength of the mutual attraction between conidia and protoperithecia decreased as genetic similarity increased.

New mutations that suppress mating-type vegetative incompatibility in Neurospora crassa

Genome ◽  
1994 ◽  
Vol 37 (2) ◽  
pp. 249-255 ◽  
Author(s):  
Tia S. Vellani ◽  
Anthony J. F. Griffiths ◽  
N. Louise Glass
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Vegetative Incompatibility ◽  
Type Locus ◽  
Regulatory Locus ◽  
Mating Type Locus ◽  
Recessive Mutations ◽  
Incompatibility Locus ◽  
New Alleles ◽  
New Mutations

The mating-type locus in the ascomycete Neurospora crassa functions as a vegetative-incompatibility locus during asexual growth such that A + a heterokaryons and A/a partial diploids are inhibited in their growth. In this study, we sought mutations that suppress mating-type associated vegetative incompatibility by using A/a partial diploids. Mutants were selected as spontaneous escapes from inhibited growth. Suppressors were identified by selecting escapes that retained the capacity to mate with both A and a strains. The escaped partial diploids were then outcrossed to remove the duplication. Seven suppressors were identified that segregated as single, recessive mutations. All seven of the suppressors were shown to be allelic to a previously obtained suppressor, tol, by segregation analyses and heterokaryon tests. This result indicates that all seven mutations are new alleles of tol and suggests that tol is a key regulatory locus in the expression of mating-type associated vegetative incompatibility in N. crassa.Key words: vegetative incompatibility, mating type, suppressors, Neurospora.

Effects of thetolmutation on allelic interactions athetloci inNeurospora crassa

Genome ◽  
1997 ◽  
Vol 40 (6) ◽  
pp. 834-840 ◽  
Author(s):  
John F. Leslie ◽  
Carl T. Yamashiro
Keyword(s):  
Neurospora Crassa ◽  
Mating Type ◽  
Vegetative Compatibility ◽  
Vegetative Incompatibility ◽  
Type Locus ◽  
Mating Type Locus ◽  
Inneurospora Crassa ◽  
Incompatibility Reactions

A mutant at the tol locus of Neurospora crassa can suppress heterokaryon (vegetative) incompatibility associated with differences at the mating-type locus. We tested the ability of this allele to suppress the vegetative incompatibility reactions that can occur when strains differ at one of nine het loci (het-C, -D, -E, -5, -6, -7, -8, -9, and -10). We found no cases in which the tol mutant suppresses a heteroallelic het locus interaction. Thus, the interaction(s) that leads to vegetative incompatibility because of differences at the mating-type locus is distinct from the interaction(s) that leads to vegetative incompatibility because of differences at any of these nine het loci.Key words: heterokaryon, mating type, vegetative compatibility.

scholarly journals Molecular Characterization of tol, a Mediator of Mating-Type-Associated Vegetative Incompatibility in Neurospora crassa

Genetics ◽  
1999 ◽  
Vol 151 (2) ◽  
pp. 545-555 ◽  
Author(s):  
Patrick Ka Tai Shiu ◽  
N Louise Glass
Keyword(s):  
Sexual Reproduction ◽  
Neurospora Crassa ◽  
Mating Type ◽  
Vegetative Growth ◽  
Point Mutations ◽  
Coiled Coil ◽  
Wild Type ◽  
Opposite Mating Type ◽  
Type Locus

Abstract The mating-type locus in the haploid filamentous fungus, Neurospora crassa, controls mating and sexual development. The fusion of reproductive structures of opposite mating type, A and a, is required to initiate sexual reproduction. However, the fusion of hyphae of opposite mating type during vegetative growth results in growth inhibition and cell death, a process that is mediated by the tol locus. Mutations in tol are recessive and suppress mating-type-associated heterokaryon incompatibility. In this study, we describe the cloning and characterization of tol. The tol gene encodes a putative 1011-amino-acid polypeptide with a coiled-coil domain and a leucine-rich repeat. Both regions are required for tol activity. Repeat-induced point mutations in tol result in mutants that are wild type during vegetative growth and sexual reproduction, but that allow opposite mating-type individuals to form a vigorous heterokaryon. Transcript analyses show that tol mRNA is present during vegetative growth but absent during a cross. These data suggest that tol transcription is repressed to allow the coexistence of opposite mating-type nuclei during the sexual reproductive phase. tol is expressed in a mat A, mat a, A/a partial diploid and in a mating-type deletion strain, indicating that MAT A-1 and MAT a-1 are not absolutely required for transcription or repression of tol. These data suggest that TOL may rather interact with MAT A-1 and/or MAT a-1 (or downstream products) to form a death-triggering complex.

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