Genome-tagged mice (GTM): two sets of genome-wide congenic strains.

An important approach for understanding complex disease risk using the mouse is to map and ultimately identify the genes conferring risk. Genes contributing to complex traits can be mapped to chromosomal regions using genome scans of large mouse crosses. Congenic strains can then be developed to fine-map a trait and to ascertain the magnitude of the genotype effect in a chromosomal region. Congenic strains are constructed by repeated backcrossing to the background strain with selection at each generation for the presence of a donor chromosomal region, a time-consuming process. One approach to accelerate this process is to construct a library of congenic strains encompassing the entire genome of one strain on the background of the other. We have employed marker-assisted breeding to construct two sets of overlapping congenic strains, called genome-tagged mice (GTMs), that span the entire mouse genome. Both congenic GTM sets contain more than 60 mouse strains, each with on average a 23-cM introgressed segment (range 8 to 58 cM). C57BL/6J was utilized as a background strain for both GTM sets with either DBA/2J or CAST/Ei as the donor strain. The background and donor strains are genetically and phenotypically divergent. The genetic basis for the phenotypic strain differences can be rapidly mapped by simply screening the GTM strains. Furthermore, the phenotype differences can be fine-mapped by crossing appropriate congenic mice to the background strain, and complex gene interactions can be investigated using combinations of these congenics.

Microsatellite marker panels for use in high-throughput genotyping of mouse crosses.

Several microsatellite genotyping panel sets have been developed that are polymorphic between C57BL/6J and CAST/Ei mice, or C57BL/6J and DBA/2J. One set of markers for each strain pair has an intermarker distance of approximately 20 cM, and a second set has an intermarker distance of 5 cM. The 20-cM set contains 105 markers for C57BL/6J x DBA/2J and 108 for C57BL/6J x CAST/Ei, divided into 13 panels. Each 5-cM set includes 350 markers arranged into 45 panels. A panel contains a number of primer pairs whose fluorescently labeled PCR products can be pooled together and separated on one lane of a polyacrylamide gel. The sets are arranged by the size of the PCR product and by the type of fluorescent dye; 5-cM sets are also arranged by chromosomal region. The 20-cM sets are most useful for full-genome scans, the 5-cM sets are useful for full-genome and/or for region-specific chromosome screens. Both sets were proven as useful tools for speed congenic development, quantitative trait loci (QTL) analysis and physical mapping. These panel sets provide a throughput of 1,536-2,304 mouse genotypes daily per one gel-based system. Whole genome scans of one animal require 13 or 48 gel lanes, with 20 cM or 5 cM density, respectively.

A genome-wide search identifies potential new susceptibility loci for Crohn's disease.

Chronic inflammatory bowel disease (IBD) presents as two major clinical forms, Crohn's disease (CD) and ulcerative colitis (UC). Genetic epidemiological studies and animal models suggest that inherited factors play significant roles in the susceptibility to both forms of IBD. From four genome-wide scans, putative susceptibility loci on chromosome 16 (IBD1 for CD), and on chromosomes 1, 3, 4, 6, 7, 10, and 12 for IBD, have been identified. Several other groups, including ours, have confirmed linkage to the loci on chromosomes 12 and 16. The aim of this study is to identify other potential susceptibility loci for CD with a genome-wide search approach. In our sample of 222 individuals from 46 families (20 Jewish and 26 non-Jewish), with a total of 65 sibpairs diagnosed with CD, we observed a novel locus with suggestive linkage [multipoint logarithm of the odds score (Mlod) > 2] at chromosome 14q11.2 (Mlod = 2.8, p = 0.0002). In addition, suggestive linkage was observed in our Jewish families at chromosome 17q21-q23 (Mlod = 2.1, p = 0.01) and chromosome 5q33-q35 (Mlod = 2.2, p = 0.0003). The syntenic regions of the latter locus are mapped within two putative loci on mouse chromosomes 11 and 18, which were identified in a mouse IBD model induced by dextran sulfate sodium (29). Our preliminary results provide potential evidence for several susceptibility loci contributing to the risk of CD. The observation of man-mouse synteny may accelerate the identification of CD susceptibility gene(s) on human chromosome 5.

Additional evidence of linkage between Crohn's disease and a putative locus on chromosome 12.

PURPOSE: The inflammatory bowel diseases (IBD), Crohn's disease (CD), and ulcerative colitis (UC) are chronic intestinal disorder of unknown etiology. Genetic factors play an important role in the pathogenesis of these diseases, but with a complex pattern of inheritance. A number of genome-wide scans have identified several putative susceptibility loci for both CD and UC, including a locus on chromosome 12 reported in a set of British families. We aim to evaluate the linkage between CD or UC and this chromosome 12 locus in an independent set of U.S. Caucasian families (36% being of Ashkenazi Jewish origin). METHODS: Microsatellite markers along chromosome 12 spaced at approximately 10 cm intervals were used to test the putative loci in CD only families (65 sib pairs from 46 families). Regions with positive linkage for CD were then tested in a panel of UC and mixed families (44 sib pairs from 29 families). Two point linkage analysis was performed with SIBPAL. Multipoint linkage analysis was carried out with MAPMAKER/SIBS. RESULTS: We observed evidence of linkage between the region on chromosome 12 and Crohn's disease, because there was a significant excess of allele sharing in CD sib pairs (pi = 0.62, p = 0.0004 from two-point linkage; and logarithm of the odds score (LOD) = 2.0 from multipoint linkage). However, we did not observe the same linkage in UC and mixed families (p = 0.48; not significant [ns]). CONCLUSION: Our data provided further evidence that the region on chromosome 12 is likely to contain a gene predisposing to CD.

Susceptibility locus for inflammatory bowel disease on chromosome 16 has a role in Crohn's disease, but not in ulcerative colitis.

In the Western world, chronic inflammatory bowel disease (IBD) presents as two major clinical forms, Crohn's disease (CD) and ulcerative colitis (UC) [Targan, S.R. and Shanahan, F. (1994). In Retford, D.C (ed.), Inflammatory Bowel Disease: From Bench to Bedside. Williams and Wilkins, Baltimore]. Genetic epidemiological studies, the occurrence of rare syndromes associated with IBD, and animal models suggest that inherited factors play significant roles in the susceptibility to both forms of IBD [Yang, H.-Y. and Rotter, J.I. (1995) In Kirsner, J.B. and Shorter, R.G. (eds). Genetic Aspects of Idiopathic Inflammatory Bowel Disease. Williams and Wilkins, Baltimore, pp.301-331]. Recently, a genome-wide search on European families with multiple affected members with CD identified a putative susceptibility locus in the centromeric region of chromosome 16 [Hugot, J.-P. et al. (1996) Nature, 379, 821-823]. We have now tested this region in an independent set of US families, confirmed that this region is likely to contain a gene predisposing to CD, and further refined the chromosomal location of this gene. Most importantly with respect to this locus, our data also seem to indicate that there is heterogeneity both within the CD group, and between the CD and UC groups with respect to this locus. The susceptibility locus appears to be involved only in non-Jewish CD sibpairs and not in our Ashkenazi Jewish CD sibpairs. Additionally, we have tested sibpairs having either only UC or both UC and CD for involvement of this locus, and have found no evidence that this region predisposes to IBD in these patients.

Seminal plasma proteins of a line of chickens selected for fertility of frozen-thawed semen and the control line.

The seminal plasma from individual ejaculates of 13 males of a line of chickens selected for increased duration of fertility of frozen-thawed semen for 7 generations and 12 males of the control line were analyzed for proteins. The mean +/- SE total seminal plasma protein of the selected and control lines were 8.74 +/- .55 micrograms and 7.52 +/- .92 micrograms/microliter seminal plasma (P greater than .05). Crossed immunoelectrophoretic analysis of the seminal plasma generated, in general, 11 precipitant peaks in both lines. The protein corresponding to peak 11 of the electrophoretic pattern was significantly (P less than .01) higher in the selected line than in the control line, the mean +/- SE being 1.80 +/- .21 and .85 +/- .19 micrograms/microliter seminal plasma.