A high-density single nucleotide polymorphism map for Neurospora crassa.
We report the discovery and validation of a set of single nucleotide polymorphisms (SNPs) between the reference Neurospora crassa strain Oak Ridge and the Mauriceville strain (FGSC 2555), of sufficient density to allow fine mapping of most loci. Sequencing of Mauriceville cDNAs and alignment to the completed genomic sequence of the Oak Ridge strain identified 19,087 putative SNPs. Of these, a subset was validated by cleaved amplified polymorphic sequence (CAPS), a simple and robust PCR-based assay that reliably distinguishes between SNP alleles. Experimental confirmation resulted in the development of 250 CAPS markers distributed evenly over the genome. To demonstrate the applicability of this map, we used bulked segregant analysis followed by interval mapping to locate the csp-1 mutation to a narrow region on LGI. Subsequently, we refined mapping resolution to 74 kbp by developing additional markers, resequenced the candidate gene, NCU02713.3, in the mutant background, and phenocopied the mutation by gene replacement in the WT strain. Together, these techniques demonstrate a generally applicable and straightforward approach for the isolation of novel genes from existing mutants. Data on both putative and validated SNPs are deposited in a customized public database at the Broad Institute, which encourages augmentation by community users.
Lambreghts
Randy
R
Department of Environmental and Biomolecular Systems, Oregon Health & Science University, Beaverton, Oregon 97006, USA.
Shi
Mi
M
Belden
William J
WJ
Decaprio
David
D
Park
Danny
D
Henn
Matthew R
MR
Galagan
James E
JE
Bastürkmen
Meray
M
Birren
Bruce W
BW
Sachs
Matthew S
MS
Dunlap
Jay C
JC
Loros
Jennifer J
JJ
D002874:Chromosome Mapping; D004271:DNA, Fungal / Q000235:genetics; D030561:Databases, Nucleic Acid; D020224:Expressed Sequence Tags; D005800:Genes, Fungal; D005819:Genetic Markers; D009154:Mutation; D009492:Neurospora crassa / Q000145:classification / Q000235:genetics*; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D011995:Recombination, Genetic; D013045:Species Specificity
D004271:DNA, Fungal; D005819:Genetic Markers