Reference
The Collaborative Cross (CC) is a large (~1,000 line) panel of recombinant inbred (RI) mouse strains being developed through a community effort (Churchill et al. 2004). The CC combines the genomes of eight genetically diverse founder strains - A/J, C57BL/6J, 129S1/SvImJ, NOD/ShiLtJ, NZO/HlLtJ, CAST/EiJ, PWK/PhJ, and WSB/EiJ - to capture nearly 90% of the known variation present in laboratory mice. CC strains are derived using a unique funnel breeding scheme. Once inbred, the RI CC lines can be used to generate thousands of potential 'outbred' but completely reproducible genomes through the generation of recombinant inbred crosses (RIX). The designation 'PreCC' is used to describe a mapping population of CC mice that is still at incipient stages of inbreeding.
CTC (2004), Churchill, G. A., et al.. The Collaborative Cross, a community resource for the genetic analysis of complex traits. Nat Genet. 36, 1133-7.
Experiment
Familial adenomatous polyposis is an inherited genetic disease, characterized by colorectal polyps. It is caused by inactivating mutations in the Adenomatous polyposis coli (Apc) gene. Mice carrying a nonsense mutation in the Apc gene at R850, which is designated ApcMin/+ (Multiple intestinal neoplasia), develop intestinal adenomas. Several genetic modifier loci of Min (Mom) were previously mapped, but so far, most of the underlying genes have not been identified. To identify novel modifier loci associated with ApcMin/+, the authors performed quantitative trait loci (QTL) analysis for polyp development using 49 F1 crosses between different Collaborative Cross (CC) lines and C57BL/6 J-ApcMin/+ mice. C57BL/6 J-ApcMin/+ males were mated with females from 49 CC/Tau lines. In total, 957 F1 mice were produced by a cross of females from 49 CC/Tau lines to C57B/6 J-ApcMin/+ males and after PCR analysis for Min genotype, 402 F1 CC/Tau-C57BL/6-ApcMin/+ (CC/Tau-B/6-ApcMin/+) mice were identified and included in the study for further assessment and analysis. F1 offspring were terminated at 23 weeks and polyp counts from three sub-regions (SB1-3) of small intestinal and colon were recorded.
The number of polyps in all these sub-regions and colon varied significantly between the different CC/Tau lines.
High molecular genomic DNA of the CC/Tau lines were initially genotyped with the mouse diversity array (MDA), which consists of 620,000 SNPs and re-genotyped by mouse universal genotype array (MUGA - 7,500 markers) and eventually with MegaMuga (77,800 markers) SNP arrays to confirm their genotype status. Data analysis was performed using the statistical software R (R Development Core Team 2009), including the R package HAPPY.HBREM [23]. The authors removed SNPs with heterozygous or missing genotypes in the 8 CC/Tau founders, or were not in common between the arrays, leaving 170,935 SNPs. The SNPs were mapped onto NCBI Build 37 of the mouse genome. The authors reconstructed the genome mosaic of each CC/Tau line in terms of the eight CC founders using a hidden Markov Model HAPPY [23].
The presence of a QTL at a given locus was tested using the probabilities of descent from each founder calculated through HAPPY and testing for association between the founder haplotype at each locus and the median polyp count within each CC line, using multiple linear regression. Sex was included as a covariate. QTL effect sizes were estimated as the proportion of the log-likelihood explained by the locus effects at the QTL. Genome-wide significance was estimated by permutation, where the CC line labels were permuted between the phenotypes.
At 90% genome-wide significance, the authors mapped 13 novel QTL for variation in polyp number, with distinct QTL associated with each intestinal sub-region (genome coordinates relative to GRCm38/mm10):
Mom22 (modifier of Min 22, small bowel 1) maps to Chr 3: 9.902 - 19.627 Mb with a peak LOD score of 4.43 at 13.839 Mb. For Mom22 there were slight positive effects on poly counts for CAST/EiJ, NZO/HlLtJ, 129S1/SvImJ strains and minimal negative effects for A/J, C57BL/6J, NOD/ShiLtJ, and PWK/PhJ.
Mom23 (modifier of Min 23, small bowel 1) maps to Chr 12: 102.018 - 118.857 Mb with a peak LOD score of 4.43 at 111.371 Mb. For Mom23 all founder strains have positive effects except A/J and C57BL/6J.
Mom24 (modifier of Min 24, small bowel 2) maps to Chr 10: 8.902 - 28.471 Mb with a peak LOD score of 4.11 at 18.805 Mb. For Mom24 all the founder strains contributed a positive effect on polyps count (i.e. this QTL involved a contrast between WSB/EiJ vs the rest).
Mom25 (modifier of Min 25, small bowel 2) maps to Chr 16: 45.522 - 63.132 Mb with a peak LOD score of 4.11 at 53.511 Mb. For Mom25 all the founder strains except PWK/PhJ contributed positive effects.
Mom26 (modifier of Min 26, small bowel 2) maps to Chr 16: 68.716 - 78.148 Mb with a peak LOD score of 4.11 at 73.216 Mb. For Mom26 all the founder strains except PWK/PhJ contributed positive effects.
Mom27 (modifier of Min 27, small bowel 3) maps to Chr 6: 138.051 - 147.806 Mb with a peak LOD score of 4.20 at 146.203 Mb. For Mom27 positive effects were seen in all the founder strains, except CAST/EiJ and 129S1/SvImJ.
Mom28 (modifier of Min 28, small bowel 3) maps to Chr 12: 109.825 - 116.663 Mb with a peak LOD score of 4.20 at 113.449 Mb. For Mom28 only A/J and C57BL/6J strain had a minor negative effect on polyps count.
Mom29 (modifier of Min 29, small bowel 3) maps to Chr 9: 32.557 - 42.557 Mb with a peak LOD score of 4.20 at 37.552 Mb. For Mom29 C57BL/6J, CAST/EiJ, and 129S1/SvImJ had a negative effect.
Mom30 (modifier of Min 30, small bowel 3) maps to Chr 10: 8.950 - 27.582 Mb with a peak LOD score of 4.20 at 18.805 Mb. For Mom30 all the founder strains contributed a positive effect on polyps count (i.e. this QTL involved a contrast between WSB/EiJ vs the rest).
Mom31 (modifier of Min 31, colon) maps to Chr 6: 34.720 - 38.392 Mb with a peak LOD score of 4.19 at 35.915 Mb. For Mom31 C57BL/6J, CAST/EiJ, and 129S1/SvImJ had a negative effect.
Mom32 (modifier of Min 32, total polyps) maps to Chr 12: 109.525 - 113.920 Mb with a peak LOD score of 4.23 at 111.636 Mb. For Mom32 all founder strains have positive effects except A/J and C57BL/6J.
Mom33 (modifier of Min 33, total polyps) maps to Chr 16: 44.055 - 63.294 Mb with a peak LOD score of 4.23 at 53.489 Mb. For Mom33 all the founder strains except PWK/PhJ contributed positive effects.
Mom34 (modifier of Min 34, total polyps) maps to Chr 16: 65.848 - 83.013 Mb with a peak LOD score of 4.23 at 73.556 Mb. For Mom34 all the founder strains except PWK/PhJ contributed positive effects.
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