title: Mutation discovery in the mouse using genetically guided array capture and resequencing.
abstract: Forward genetics (phenotype-driven approaches) remain the primary source for allelic variants in the mouse. Unfortunately, the gap between observable phenotype and causative genotype limits the widespread use of spontaneous and induced mouse mutants. As alternatives to traditional positional cloning and mutation detection approaches, sequence capture and next-generation sequencing technologies can be used to rapidly sequence subsets of the genome. Application of these technologies to mutation detection efforts in the mouse has the potential to significantly reduce the time and resources required for mutation identification by abrogating the need for high-resolution genetic mapping, long-range PCR, and sequencing of individual PCR amplimers. As proof of principle, we used array-based sequence capture and pyrosequencing to sequence an allelic series from the classically defined Kit locus (approximately 200 kb) from each of five noncomplementing Kit mutants (one known allele and four unknown alleles) and have successfully identified and validated a nonsynonymous coding mutation for each allele. These data represent the first documentation and validation that these new technologies can be used to efficiently discover causative mutations. Importantly, these data also provide a specific methodological foundation for the development of large-scale mutation detection efforts in the laboratory mouse.
authors.affiliation: Roche NimbleGen, 500 South Rosa Road, Madison, WI 53719, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: D'Ascenzo
authors.forename: Carl
authors.initials: C
authors.lastname: Meacham
authors.forename: Jacob
authors.initials: J
authors.lastname: Kitzman
authors.forename: Christina
authors.initials: C
authors.lastname: Middle
authors.forename: Jim
authors.initials: J
authors.lastname: Knight
authors.forename: Roger
authors.initials: R
authors.lastname: Winer
authors.forename: Miroslav
authors.initials: M
authors.lastname: Kukricar
authors.forename: Todd
authors.initials: T
authors.lastname: Richmond
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Albert
authors.forename: Anne
authors.initials: A
authors.lastname: Czechanski
authors.forename: Leah Rae
authors.initials: LR
authors.lastname: Donahue
authors.forename: Jason
authors.initials: J
authors.lastname: Affourtit
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Jeddeloh
authors.forename: Laura
authors.initials: L
authors.lastname: Reinholdt
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004252:DNA Mutational Analysis / Q000379:methods*; D005260:Female; D008297:Male; D051379:Mice / Q000235:genetics*; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008969:Molecular Sequence Data; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016415:Sequence Alignment