Isolation of chromosome-specific paints from high-resolution flow karyotypes of the sheep (Ovis aries).
High-resolution bivariate flow karyotypes were obtained using fibroblast cell lines from a sheep with a normal karyotype (2n = 54), from sheep carrying Robertsonian translocation chromosomes and from sheep-hamster somatic cell hybrids. By taking advantage of the presence of chromosome polymorphisms, translocation chromosomes and sheep-hamster somatic cell hybrids, all sheep chromosomes were isolated by flow sorting. Chromosome-specific paints were generated from each sorted peak using degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR). The sheep chromosome present in each peak was identified by chromosome-specific microsatellite analysis of the DOP-PCR products and fluorescence in situ hybridization (FISH) onto DAPI-banded sheep metaphase chromosomes. The chromosome-specific DNA obtained in this study can be used for the production of genomic libraries and as a resource for mapping randomly cloned DNA sequences that will greatly aid the construction of genetic and physical maps in the sheep. The chromosome-specific paints will facilitate chromosome identification and contribute to the study of karyotype evolution in the sheep and related species.
Burkin
D J
DJ
Pathology Department, University of Cambridge, UK. maf12@mole.bio.cam.ac.uk
O'Brien
P C
PC
Broad
T E
TE
Hill
D F
DF
Jones
C A
CA
Wienberg
J
J
Ferguson-Smith
M A
MA
D000818:Animals; D002460:Cell Line; D002469:Cell Separation; D006224:Cricetinae; D015342:DNA Probes; D005347:Fibroblasts; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D006822:Hybrid Cells; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007211:Indoles; D007621:Karyotyping; D008297:Male; D018895:Microsatellite Repeats / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D012756:Sheep / Q000235:genetics*; D014178:Translocation, Genetic
D015342:DNA Probes; D005456:Fluorescent Dyes; D007211:Indoles; C007293:DAPI