Chromosomal localization and molecular characterization of 53 cosmid-derived bovine microsatellites

Abstract

Gene mapping in cattle has progressed rapidly in recent years largely owing to the introduction of powerful genetic markers, such as the microsatellites, and through advances in physical mapping techniques such as synteny mapping and fluorescence in situ hybridization (FISH). Microsatellite markers are often not physically mapped because they are generally isolated from small insert plasmid libraries, which makes their chromosomal localization inefficient. In this report we describe the FISH mapping of a large group of cosmid-derived bovine microsatellite markers, as our contribution to the European mapping initiative, BovMap. One objective of BovMap is to develop a set of anchored loci for the cattle genome map. Two cosmid libraries were screened with probes corresponding to the (AC)n microsatellite motif. Positive clones were mapped by FISH, and then a subset was further analyzed by sequencing the region flanking the microsatellite repeat. In total, 58 clones were hybridized with chromosomes and identified loci on 22 of the 31 different bovine chromosomes. Three clones contained satellite DNA. Two or more markers were placed on 12 chromosomes. Sequencing of the microsatellites and flanking regions was performed directly from 43 cosmids, as previously reported (Ferretti et al. Anim. Genet. 25, 209-214, 1994). Primers were developed for 39 markers and used to describe the polymorphism associated with the corresponding loci.

Cosmid-derived markers anchoring the bovine genetic map to the physical map

“…No data are presented for Chrs 13 or X, which only have two markers mapped at adjoining positions. In total, intervals could be considered for 20 chro- X98441 / (Williams et al 1996a) X93164/ (Williams et al 1996b) X97565 / X85053 / Mezzelani et al 1995) Z27071 / X85070 / X85052 / X89258 / X73947 / X85046 / X85068 / 7 Z73084 / X86815 / X86811 / X03162 / (Anastassiadis et al 1996) Z27077 / X85069/ Mezzelani et al 1995) X74174/ (Eggen et al 1994) X98445 / (Williams et al 1996a) X89255 / X74201 / Z27072 / X73945 / X98442 / (Williams et al 1996a) X85072 / X81349 / (Thieven et al 1995 X71495 / (Vaiman et al 1993) Z27076 / X85051 / X89249 / X95067 / Z73082 / (Williams et al 1996e) X85081 / Z22740 / (Solinas Toldo et al 1993) X98444/ (Williams et al 1996a) X85055/ U47616 / (Martin-Burriel et al 1996a) X85050 / X81350 / (Thieven et al 1995) X85071/ X85062 / U31002 / X85059 / Z22744 / (Solinas Toldo et al 1993) (Eggen 1992) X85060 / X89253 / X85054 / X85049/ X73944 / X95068 / U59512 / (Olsaker et al 1996c) X95069 / X85053 / X89260 / U51935 / X85074 / U59511 / U51932 / X85048 / X85058 / Z27074 / Z27073 / X98440 / (Williams et al 1996a) U51933/ mosomes, covering 40.6% of the whole genome (Table 2). In some cases, the size of the interval used was increased by including genes for which physical and linkage mapping data were available.…”

“…For the majority of clones (that is, IDVGA and ETH clones), QFQ-banded chromosome preparations were subjected to competitive in sitn suppression hybridization (Lichter et al 1990) and then hybridzed with biotinylated probes. FITC signal detection was performed with a computercontrolled CCD camera device (Photometrics, Tucson, Ariz.), as described by Solinas Toldo et al (1993) and Mezzelani et al (1995). Chromosomal band assignments were obtained by determining the fractional length (FLcen) with respect to the proximal border of the chromosome and superimposing the value to the idiogram of the standard chromosomes (Popescu et al 1996).…”

“…This has been achieved in most cases by combining the localization of large-insert clones to chromosomes by FISH, with the PCR amplification of microsatellites derived from the clones on hybrid cell lines. Obtaining highly polymorphic markers from cosmid and phage clones is a very efficient way of placing markers on both the genetic and physical maps (Solinas Toldo et al 1993;Ellegren et al 1994a;Eggen and Fries 1995;Mezzelani et al 1995;Thieven et al 1996). This approach was adopted by the BovMap group as the tool of choice to provide physical anchorage to the developing bovine linkage map, and the results are presented in this paper.…”

Anchoring of bovine chromosomes 4, 6, 7, 10, and 14 linkage group telomeric ends via FISH analysis of lambda clones

“…The usefulness of the genetic map in identifying ETL is a function of the overall coverage of the genome and the integration of the genetic linkage map with the cytogenetic (physical) map. A major goal of map construction has thus been the generation of polymorphic markers physically localized to regular intervals along the chromosome and at the extremes of the linkage groups, to use for scanning the genome of populations segregating phenotypes of interest (Fries 1993;SolinasToldo et al 1993;Beattie 1994;Ellegren et al 1994;Ferretti et al 1994;Smith et al 1995;Hawkins et al 1995;Mezzelani et al 1995).…”

“…The difficulty of physical localization of the small inserts typically used to identify and sequence ms Barendese et al 1994;Stone et al 1995) has limited the number of linked, highly polymorphic markers that have been physically assigned in the bovine genome by in situ techniques (Mezzelani et al 1995;reviewed in Eggen and Fries, 1995). This lack of anchors makes genome coverage estimates of the genetic linkage map difficult.…”