Targeted mutagenesis in mice, a powerful tool for the analysis of gene function and human disease, makes extensive use of 129 mouse substrains. Although all are named 129, we document that outcrossing of these substrains, both deliberate and accidental, has lead to extensive genetic variability among substrains and embryonic stem cells derived from them. This clearer understanding of 129 substrain variability allows consideration of its negative impact on targeting technology, including: homologous recombination frequencies, preparation of inbred animals, and availability of appropriate controls. Based on these considerations we suggest a number of recommendations for future experimental design.

A set of 1638 informative SNP markers easily assayed by the Amplifluor genotyping system were tested in 102 mouse strains, including the majority of the common and wild-derived inbred strains available from The Jackson Laboratory. Selected from publicly available databases, the markers are on average ∼1.5 Mb apart and, whenever possible, represent the rare allele in at least two strains. Amplifluor assays were developed for each marker and performed on two independent DNA samples from each strain. The mean number of polymorphisms between strains was 608±136 SD. Several tests indicate that the markers provide an effective system for performing genome scans and quantitative trait loci analyses in all but the most closely related strains. Additionally, the markers revealed several subtle differences between closely related mouse strains, including the groups of several 129, BALB, C3H, C57, and DBA strains, and a group of wild-derived inbred strains representing several Mus musculus subspecies. Applying a neighbor-joining method to the data, we constructed a mouse strain family tree, which in most cases confirmed existing genealogies.

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SUMMARY:New Zealand Obese (NZO)/HlLt male mice exhibit a polygenic obesity and approximately 50% develop type 2 diabetes. This strain is known to produce a variety of autoantibodies, including autoantibodies to the insulin receptor. Because of their relatedness to the autoimmune-predisposed New Zealand Black (NZB) and New Zealand White (NZW) inbred strains, we compared NZO to its two related strains for shared hematologic and immunologic characteristics. Comparison of the three strains by serotyping and genotyping methods indicated that NZO shared with NZW the rare (recombinant) H2 z haplotype at the major histocompatibility complex. Similar to the NZB and NZW strains, spleens from NZO mice contained increased numbers of CD19 ϩ CD43 ϩ IgM ϩ B-1 B cells, a phenotype associated with natural autoantibody production. NZO mice developed a progressive microcytic anemia that was distinguished from NZB hemolytic anemia by absence of demonstrable antierythrocyte antibodies in the former. Outcross of NZO females with NZB males accelerated development of obesity and diabetes in F1 males. NZO males made B-lymphocyte-deficient by a disrupted immunoglobulin heavy chain gene did not become diabetic. These results suggest that NZO mice should be useful to investigators interested in studying the genetic contributions to autoimmunity made by the related NZW and NZB strains. Further, these results, combined with the pancreatic histopathology contained in the companion manuscript, suggest that B lymphocytes may be important contributors to diabetes pathogenesis in the NZO mouse. (Lab Invest 2002, 82:833-842).

The success of high resolution genetic mapping of disease predisposition and quantitative trait loci in humans and experimental animals depends on the positions of key crossover events around the gene of interest. In mammals, the majority of recombination occurs at highly delimited 1–2 kb long sites known as recombination hotspots, whose locations and activities are distributed unevenly along the chromosomes and are tightly regulated in a sex specific manner. The factors determining the location of hotspots started to emerge with the finding of PRDM9 as a major hotspot regulator in mammals, however, additional factors modulating hotspot activity and sex specificity are yet to be defined. To address this limitation, we have collected and mapped the locations of 4829 crossover events occurring on mouse chromosome 11 in 5858 meioses of male and female reciprocal F1 hybrids of C57BL/6J and CAST/EiJ mice. This chromosome was chosen for its medium size and high gene density and provided a comparison with our previous analysis of recombination on the longest mouse chromosome 1. Crossovers were mapped to an average resolution of 127 kb, and thirteen hotspots were mapped to <8 kb. Most crossovers occurred in a small number of the most active hotspots. Females had higher recombination rate than males as a consequence of differences in crossover interference and regional variation of sex specific rates along the chromosome. Comparison with chromosome 1 showed that recombination events tend to be positioned in similar fashion along the centromere-telomere axis but independently of the local gene density. It appears that mammalian recombination is regulated on at least three levels, chromosome-wide, regional, and at individual hotspots, and these regulation levels are influenced by sex and genetic background but not by gene content.

McManus advanced a genetic hypothesis to explain differences of lateralization between HI and LO lines of mice selectively bred for degree of handedness. It states that lateralization is a function of heterozygosity. Specifically it predicts that (a) the HI line will be more heterozygous than the LO line and (b) populations with a greater average heterozygosity (AH) will be more strongly lateralized. Both genetic and behavioral predictions were tested here. Results using coat color and biochemical variants show that AH in the HI line is somewhat less (not greater) than that in the LO line. The handedness of HET control mice and HI by LO reciprocal hybrids, where AH is greater than that of the HI line, exhibits lessened (not greater) lateralization. Results reject the heterozygosity hypothesis. A model for the inheritance of human handedness that accounts for difficulty in detecting heritable differences in degree of asymmetry is presented.

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To search for host genes for resistance/susceptibility to cancer metastasis, mutation analysis was employed. Ten putative mutants of resistance to lymphoma EL4 and four putative mutants of resistance to sarcoma MCA/77-23 of C57BL/6J (B6) mice were produced. These mutants were designated S (for "survivor") mutants; they do not reject parental strain B6 skin grafts. S-mutants resist moderate tumor cell doses: TD50 values in them were increased by a factor of 12 to 600. Genetic linkage tests showed that five S-mutants were linked to mouse major histocompatibility complex (H-2) and five other S-mutants were not linked to this locus. A group of H-2-linked S-mutants resisting EL4 and a mutant, S-87/2, resisting MCA/77-23 were tested for resistance to spontaneous metastases of the same two tumors, EL4 and MCA/77-23. Two of the mutants, S-31 (lymphoma-resisting) and S-87/2 (sarcoma-resisting), were shown to carry mutations of mouse gene(s) for resistance to tumor metastases. In both of these mutants resistance to the original tumor transplant coexisted with highly increased susceptibility to metastasis. These mutants are a new tool to study genes for resistance to cancer metastasis and of mechanism of resistance controlled by each individual gene.