Skip to main content
Log in

Mutation discovery in the mouse using genetically guided array capture and resequencing

Mammalian Genome Aims and scope Submit manuscript

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 (~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.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

References

  • Albert TJ, Molla MN, Muzny DM, Nazareth L, Wheeler D et al (2007) Direct selection of human genomic loci by microarray hybridization. Nat Methods 4:903–905

    Article  PubMed  CAS  Google Scholar 

  • Bernstein A, Chabot B, Dubreuil P, Reith A, Nocka K et al (1990) The mouse W/c-kit locus. Ciba Found Symp 148:158–166, discussion 166–172

    PubMed  CAS  Google Scholar 

  • de Aberle SB (1927) A study of hereditary anemia of the mouse. Am J Anat 40:219–249

    Article  Google Scholar 

  • Geissler EN, McFarland EC, Russell ES (1981) Analysis of pleiotropism at the dominant white-spotting (W) locus of the house mouse: a description of ten new W alleles. Genetics 97:337–361

    PubMed  CAS  Google Scholar 

  • Geissler EN, Ryan MA, Housman DE (1988) The dominant-white spotting (W) locus of the mouse encodes the c-kit proto-oncogene. Cell 55:185–192

    Article  PubMed  CAS  Google Scholar 

  • Hodges E, Xuan Z, Balija V, Kramer M, Molla MN et al (2007) Genome-wide in situ exon capture for selective resequencing. Nat Genet 39:1522–1527

    Article  PubMed  CAS  Google Scholar 

  • Keays DA, Clark TG, Flint J (2006) Estimating the number of coding mutations in genotypic- and phenotypic-driven N-ethyl-N-nitrosourea (ENU) screens. Mamm Genome 17:230–238

    Article  PubMed  CAS  Google Scholar 

  • Little CC, Cloudman AM (1937) The occurrence of a dominant spotting mutation in the house mouse. Proc Natl Acad Sci USA 23:535–537

    Article  PubMed  CAS  Google Scholar 

  • Nocka K, Majumder S, Chabot B, Ray P, Cervone M et al (1989) Expression of c-kit gene products in known cellular targets of W mutations in normal and W mutant mice–evidence for an impaired c-kit kinase in mutant mice. Genes Dev 3:816–826

    Article  PubMed  CAS  Google Scholar 

  • Nocka K, Tan JC, Chiu E, Chu TY, Ray P et al (1990) Molecular bases of dominant negative and loss of function mutations at the murine c-kit/white spotting locus: W 37, W v, W 41 and W. EMBO J 9:1805–1813

    PubMed  CAS  Google Scholar 

  • Okou DT, Steinberg KM, Middle C, Cutler DJ, Albert TJ et al (2007) Microarray-based genomic selection for high-throughput resequencing. Nat Methods 4:907–909

    Article  PubMed  CAS  Google Scholar 

  • Quwailid MM, Hugill A, Dear N, Vizor L, Wells S et al (2004) A gene-driven ENU-based approach to generating an allelic series in any gene. Mamm Genome 15:585–591

    Article  PubMed  CAS  Google Scholar 

  • Reith AD, Rottapel R, Giddens E, Brady C, Forrester L et al (1990) W mutant mice with mild or severe developmental defects contain distinct point mutations in the kinase domain of the c-kit receptor. Genes Dev 4:390–400

    Article  PubMed  CAS  Google Scholar 

  • Schlager G, Dickie MM (1967) Spontaneous mutations and mutation rates in the house mouse. Genetics 57:319–330

    PubMed  CAS  Google Scholar 

  • She X, Cheng Z, Zollner S, Church DM, Eichler EE (2008) Mouse segmental duplication and copy number variation. Nat Genet 40:909–914

    Article  PubMed  CAS  Google Scholar 

  • Sweet HO, Cook S, Davisson MT (1990) The W allele W <73J>. Mouse Genome 86:239

    Google Scholar 

  • Thomas PD, Kejariwal A, Guo N, Mi H, Campbell MJ et al (2006) Applications for protein sequence-function evolution data: mRNA/protein expression analysis and coding SNP scoring tools. Nucleic Acids Res 34:W645–W650

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

We thank H. Halvensleben, C. Erickson, L. Freeberg, T Millard, and L. Lincoln for providing technology and logistical support operations at Roche NimbleGen; C. Turcotte and the 454 Sequencing center for library construction and sequencing support; X. Zheng and L. Dannenberg for providing project coordination support; and C. Birkenmeier and M. Berry for providing pedigree and historical data from the Kit alleles used in this study (The Jackson Laboratory). This work was supported by Roche Applied Sciences and a Cancer Center Core Grant to The Jackson Laboratory (CA34196).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Laura Reinholdt.

Additional information

M. D’Ascenzo and C. Meacham contributed equally to this work.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOC 50 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

D’Ascenzo, M., Meacham, C., Kitzman, J. et al. Mutation discovery in the mouse using genetically guided array capture and resequencing. Mamm Genome 20, 424–436 (2009). https://doi.org/10.1007/s00335-009-9200-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00335-009-9200-y

Keywords

Navigation