Isolation of rat chromosome-specific paint probes by bivariate flow sorting followed by degenerate oligonucleotide primed-PCR.
The flow karyotype of rat chromosomes was determined by dual beam flow cytometry. Eighteen fractions were sorted and subsequently amplified by degenerate oligonucleotide primed-PCR as described by Telenius et al. (1992a, 1992b). The PCR products were labeled and used as probes for fluorescence in situ hybridization on rat fibroblast metaphases. The amplified chromosomes were detectable as bright chromosome paints and in most cases the signal was evenly distributed along the whole chromosome except for the centromeric region in half of the chromosomes. The results show that chromosomes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 19, 20, X and Y can be sorted as individual fractions, whereas chromosomes 11, 13, 14, 15 and chromosomes 16, 17 and 18 are clustered together in the flow karyotype.
Hoebee
B
B
Laboratory of Carcinogenesis and Mutagenesis, National Institute of Public Health and Environmental Protection, Bilthoven, The Netherlands.
de Stoppelaar
J M
JM
Suijkerbuijk
R F
RF
Monard
S
S
D000818:Animals; D002478:Cells, Cultured; D002874:Chromosome Mapping; D017931:DNA Primers; D015342:DNA Probes; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D005434:Flow Cytometry; D017404:In Situ Hybridization, Fluorescence; D007621:Karyotyping; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods; D051381:Rats; D017208:Rats, Wistar / Q000235:genetics*; D014960:X Chromosome; D014998:Y Chromosome
D017931:DNA Primers; D015342:DNA Probes