Abstract
BackgroundThe development of powerful new methods for DNA sequencing enable the discovery of sequence variants, their utilization for the mapping of mutant loci, and the identification of causal variants in a single step. We have applied this approach for the analysis of ENU-mutagenized mice maintained on an inbred background.ResultsWe ascertained ENU-induced variants in four different phenotypically mutant lines. These were then used as informative markers for positional cloning of the mutated genes. We tested both whole genome (WGS) and whole exome (WES) datasets.ConclusionBoth approaches were successful as a means to localize a region of homozygosity, as well as identifying mutations of candidate genes, which could be individually assessed. As expected, the WGS strategy was more reliable, since many more ENU-induced variants were ascertained.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2173-1) contains supplementary material, which is available to authorized users.
Highlights
The development of powerful new methods for DNA sequencing enable the discovery of sequence variants, their utilization for the mapping of mutant loci, and the identification of causal variants in a single step
We have investigated if analysis of ENU-induced sequence variants in a pooled sample of inbred mutants using WGS could reliably identify a causal mutation associated with a disease-causing phenotype
Generation 1 (G1) males were obtained from C57BL/6 J (B6) males treated with ENU that were crossed with wild-type B6 females
Summary
The development of powerful new methods for DNA sequencing enable the discovery of sequence variants, their utilization for the mapping of mutant loci, and the identification of causal variants in a single step We have applied this approach for the analysis of ENU-mutagenized mice maintained on an inbred background. Phenotype-driven screens in mice treated with chemical mutagen N-ethyl-N-nitrosurea have proven to be a powerful means to facilitate gene discovery for a wide variety of biological processes An advantage of this method is the creation of point mutations throughout the genome in an unbiased fashion. The process of gene mapping in mice is done by crossing the inbred line on which the mutation was generated with a different inbred line, and following the segregation of the mutant phenotype with known informative genetic markers This requires a two-step analysis: genetic mapping to localize the region carrying the mutation (done presently by analysis of strain-specific single nucleotide polymorphism (SNPs)), followed by sequence analysis to detect the causal variant. WES has been used with success to find causal mutations in mice [6, 7]
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