Male Infertility Is Responsible for Nearly Half of the Extinction Observed in the Mouse Collaborative Cross.

The goal of the Collaborative Cross (CC) project was to generate and distribute over 1000 independent mouse recombinant inbred strains derived from eight inbred founders. With inbreeding nearly complete, we estimated the extinction rate among CC lines at a remarkable 95%, which is substantially higher than in the derivation of other mouse recombinant inbred populations. Here, we report genome-wide allele frequencies in 347 extinct CC lines. Contrary to expectations, autosomes had equal allelic contributions from the eight founders, but chromosome X had significantly lower allelic contributions from the two inbred founders with underrepresented subspecific origins (PWK/PhJ and CAST/EiJ). By comparing extinct CC lines to living CC strains, we conclude that a complex genetic architecture is driving extinction, and selection pressures are different on the autosomes and chromosome X Male infertility played a large role in extinction as 47% of extinct lines had males that were infertile. Males from extinct lines had high variability in reproductive organ size, low sperm counts, low sperm motility, and a high rate of vacuolization of seminiferous tubules. We performed QTL mapping and identified nine genomic regions associated with male fertility and reproductive phenotypes. Many of the allelic effects in the QTL were driven by the two founders with underrepresented subspecific origins, including a QTL on chromosome X for infertility that was driven by the PWK/PhJ haplotype. We also performed the first example of cross validation using complementary CC resources to verify the effect of sperm curvilinear velocity from the PWK/PhJ haplotype on chromosome 2 in an independent population across multiple generations. While selection typically constrains the examination of reproductive traits toward the more fertile alleles, the CC extinct lines provided a unique opportunity to study the genetic architecture of fertility in a widely genetically variable population. We hypothesize that incompatibilities between alleles with different subspecific origins is a key driver of infertility. These results help clarify the factors that drove strain extinction in the CC, reveal the genetic regions associated with poor fertility in the CC, and serve as a resource to further study mammalian infertility.

The Mouse Universal Genotyping Array: From Substrains to Subspecies.

Genotyping microarrays are an important resource for genetic mapping, population genetics, and monitoring of the genetic integrity of laboratory stocks. We have developed the third generation of the Mouse Universal Genotyping Array (MUGA) series, GigaMUGA, a 143,259-probe Illumina Infinium II array for the house mouse (Mus musculus). The bulk of the content of GigaMUGA is optimized for genetic mapping in the Collaborative Cross and Diversity Outbred populations, and for substrain-level identification of laboratory mice. In addition to 141,090 single nucleotide polymorphism probes, GigaMUGA contains 2006 probes for copy number concentrated in structurally polymorphic regions of the mouse genome. The performance of the array is characterized in a set of 500 high-quality reference samples spanning laboratory inbred strains, recombinant inbred lines, outbred stocks, and wild-caught mice. GigaMUGA is highly informative across a wide range of genetically diverse samples, from laboratory substrains to other Mus species. In addition to describing the content and performance of the array, we provide detailed probe-level annotation and recommendations for quality control.

Analyses of allele-specific gene expression in highly divergent mouse crosses identifies pervasive allelic imbalance.

Complex human traits are influenced by variation in regulatory DNA through mechanisms that are not fully understood. Because regulatory elements are conserved between humans and mice, a thorough annotation of cis regulatory variants in mice could aid in further characterizing these mechanisms. Here we provide a detailed portrait of mouse gene expression across multiple tissues in a three-way diallel. Greater than 80% of mouse genes have cis regulatory variation. Effects from these variants influence complex traits and usually extend to the human ortholog. Further, we estimate that at least one in every thousand SNPs creates a cis regulatory effect. We also observe two types of parent-of-origin effects, including classical imprinting and a new global allelic imbalance in expression favoring the paternal allele. We conclude that, as with humans, pervasive regulatory variation influences complex genetic traits in mice and provide a new resource toward understanding the genetic control of transcription in mammals.

Discovery of novel variants in genotyping arrays improves genotype retention and reduces ascertainment bias.

BACKGROUND: High-density genotyping arrays that measure hybridization of genomic DNA fragments to allele-specific oligonucleotide probes are widely used to genotype single nucleotide polymorphisms (SNPs) in genetic studies, including human genome-wide association studies. Hybridization intensities are converted to genotype calls by clustering algorithms that assign each sample to a genotype class at each SNP. Data for SNP probes that do not conform to the expected pattern of clustering are often discarded, contributing to ascertainment bias and resulting in lost information - as much as 50% in a recent genome-wide association study in dogs. RESULTS: We identified atypical patterns of hybridization intensities that were highly reproducible and demonstrated that these patterns represent genetic variants that were not accounted for in the design of the array platform. We characterized variable intensity oligonucleotide (VINO) probes that display such patterns and are found in all hybridization-based genotyping platforms, including those developed for human, dog, cattle, and mouse. When recognized and properly interpreted, VINOs recovered a substantial fraction of discarded probes and counteracted SNP ascertainment bias. We developed software (MouseDivGeno) that identifies VINOs and improves the accuracy of genotype calling. MouseDivGeno produced highly concordant genotype calls when compared with other methods but it uniquely identified more than 786000 VINOs in 351 mouse samples. We used whole-genome sequence from 14 mouse strains to confirm the presence of novel variants explaining 28000 VINOs in those strains. We also identified VINOs in human HapMap 3 samples, many of which were specific to an African population. Incorporating VINOs in phylogenetic analyses substantially improved the accuracy of a Mus species tree and local haplotype assignment in laboratory mouse strains. CONCLUSION: The problems of ascertainment bias and missing information due to genotyping errors are widely recognized as limiting factors in genetic studies. We have conducted the first formal analysis of the effect of novel variants on genotyping arrays, and we have shown that these variants account for a large portion of miscalled and uncalled genotypes. Genetic studies will benefit from substantial improvements in the accuracy of their results by incorporating VINOs in their analyses.