High-resolution gametic map of the sheep callipyge region: linkage heterogeneity among rams detected by sperm typing S Lien, N E Cockett, H Klungland, N Arnheim, M Georges, L Gomez-Raya Summary The callipyge locus (CLPG) causing muscular hypertrophy in domestic sheep has previously been mapped to the distal part of ovine chromosome 18. In this study, an accurate multipoint linkage map consisting of six micro- satellite markers in this chromosomal region was constructed based on the analysis of 1145 single sperm cells. The best supported order of markers was OARHH47-ILSTS54-MCM38- CSSM18-IDVGA30-BMS1561. The log odds against the second most likely order, which has a reversal of the closely linked markers CSSM18 and IDVGA30, was 5 . 026. Sperm typing can be used to examine a large number of meioses in single individuals, and therefore, was exploited to study individual variability of recombination rate in rams of different callipyge genotypes. The results revealed statistically significant linkage heterogeneity among rams (P <0 . 05) for marker interval OARHH47- CSSM18, with individual recombination frac- tions varying from 0 . 209 to 0 . 357. Keywords: callipyge locus, linkage heterogene- ity, sperm typing Introduction A heritable muscular hypertrophy associated with leanness and improved feed efficiency has been described in domestic sheep (Ovis aries). It has been proposed that an autosomal locus (callipyge; CLPG), mapped to the distal part of ovine chromosome 18, is responsible for the muscular hypertrophy condition, but the corre- sponding gene has not yet been identified (Cockett et al. 1994, 1996; Freking et al. 1998). In order to identify and characterize the poten- tially economically advantageous CLPG gene, efforts have been undertaken to generate a high density marker map in the CLPG region. Although relatively few markers have been mapped in sheep, conservation in sequence and chromosomal localization of markers across Bovidae allows about half of the bovine micro- satellites to be used in sheep (Moore et al. 1991; Cockett et al. 1994). Cattle-derived microsatel- lites that do amplify ovine DNA seem to be just as polymorphic as sheep-derived markers (Moore et al. 1991; Crawford et al. 1995). Although ovine chromosome 18 is relatively dense in markers (Crawford et al. 1995; Cockett et al. 1996; Freking et al. 1998), the maps generated are still low in resolution due to a limited number of informative meioses. Impor- tantly, the statistical power for accurately comparing frequencies of recombination suffers particularly in accuracy over small intervals because these distances are estimated from very rare events in a limited sample size. As an alternative to traditional linkage analy- sis within families, sperm typing (Li et al. 1988) offers a potentially unlimited number of meioses that can be analyzed for each indivi- dual. This method has been shown to be particularly useful for loci ordering and high- resolution recombination analysis of closely linked loci whose chromosomal localization has been only approximately defined by pedi- gree analysis (Goradia et al. 1991; Klungland et al. 1997). The technique has been success- fully applied to detecting variability in recom- bination rate in both humans (Yu et al. 1996) and cattle (Park et al. 1995; Simianer et al. 1997) and used to study meiotic recombinant hot spots (Hubert et al. 1994). The purpose of this study was to exploit the unique character- istics of sperm typing to efficiently construct an accurate multipoint linkage map covering the callipyge region on ovine chromosome 18, and test rams for linkage heterogeneity in the region. Materials and methods Material Seven rams were selected based on informa- tiveness of microsatellite markers and CLPG genotypes. All sheep were 2±4 years of age. Markers for the experiment were selected based Animal Genetics, 1999, 30, 42±46 S Lien H Klungland L Gomez-Raya Department of Ani- mal Science, Agricul- tural University of Norway, 1432 Aas, Norway N E Cockett Department of Ani- mal, Dairy and Veter- inary Sciences, Utah State University, Lo- gan, UT 84322±4700, USA N Arnheim Department of Biolo- gical Sciences, Uni- versity of Southern California, Los An- geles, CA 90089± 1340, USA M Georges Department of Genet- ics, Faculty of Veter- inary Medicine, University of Lie Áge (B43), 20 Bd de Co- lonster, 4000-Lie Áge, Belgium Correspondence: Sigbjùrn Lien. Accepted 6 November 1998 ã 1999 International Society for Animal Genetics 42