Abstract
The bivariate analysis by flow cytometry of human chromosome suspensions stained with the two fluorochromes, Hoechst 33258 (specificity for AT-rich DNA) and Chromomycin A3 (specificity for GC-rich DNA) enables the majority of the chromosome types to be resolved and sorted (Gray et. al. 1979). In the flow cytometer, chromosomes pass sequentially through two spatially separated laser beams, the first operated using the UV lines (351–364 nm) to excite Hoechst fluorescence and the second operated at 457.9 nm to excite Chromomycin fluorescence. The fluorescence intensity of each chromosome is measured for both dyes independently and correlated on the bivariate flow karyotype where chromosome types are resolved by DNA content and base pair ratio (see Fig. 1).
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
References
Arai N, Nomura D, Villaret D, DeWaal Malefijt R, Seiki M, Yoshida M, Minoshima S, Fukuyama R, Maekawa M, Kudoh J, Shimizu N, Yokota K, Abe E, Yokota T, Takebe Y, Arai K (1989) Complete nucleotide sequence of the chromosomal gene for human IL-4 and its expression J Immunol 142: 274–282
Bernheim A, Metezeau P, Guellaen G, Fellous M, Goldberg ME, Berger R (1983) Direct hybridization of sorted human chromosomes: Localization of the Y chromosome on the flow karyotype Proc Natl Acad Sci USA 80: 7571–7575
Carter NP, Albertson DG, Smith SJ, Rhodes DA, Cox TM (1991) Assignment of the human ferrochelatase gene (FECH) and a locus for protoporphyria to chromosomes 18q22 Genomics 11: 1152–1154
Carter NP, Ferguson-Smith MA, Perryman MT, Telenius H, Pelmear AH, Leversha MA, Glancy MT, Wood SL, Cook K, Dyson HM, Ferguson-Smith ME, Willatt LR (1992) Reverse chromosome painting: a method for the rapid analysis of aberrant chromosomes in clinical cytogenetics J Med Genet (in press)
Carter NP, Hampson RM, Harris RM, Yates JRW, Ferguson-Smith MA (1990) Gene mapping using flowasorted chromosomes Proc Royal Microscopical Soc 1: 511–514
Carter NP, Ferguson-Smith ME, Affara NA, Briggs H, Ferguson-Smith MA (1990) Study of chromosome abnormality in XX males using bivariate flow karyotype analysis and flow sorted dot blots Cytometry 11: 202–207
Collard JG, de Boer PAJ, Janssen JWG, Schijven JF, de Jong B (1985) Gene mapping by chromosome spot hybridization Cytometry 6: 179–185
Connor JM, Ferguson-Smith MA (1987) Essential Medical Genetics Blackwell Scientific Publications
Cotter F, Nasipuri S, Lam G, Young BD (1989) Gene mapping by enzymatic amplification from flow-sorted chromosomes Genomics 5: 470–474
Gray JW, Langlois RG, Carrano AV, Burkhart-Schultz K, Van Dilla MA (1979) High resolution chromosome analysis: one and two parameter flow cytometry Chromosoma 73: 9–27
Hulten MA, Gould CP, Goldman ASH, Waters JJ (1991) Chromosome in situ suppression hybridisation in clinical cytogenetics J Med Genet 28: 577–582
Lebo RV, Bruce BD (1987) Gene mapping with sorted chromosomes Methods Enzymol 151: 292–313
Lebo RV, Gorin F, Fletterick RJ, Kao F-K, Cheung M-C, Bruce BD, Kan YW (1984) High-resolution chromosome sorting, DNA spot-blot analysis assign McArdle’s syndrome to Chromosome 11 Science 225: 57–59
Neckelmann N, Warner CK, Chung A, Kudoh J, Minoshima S, Fukuyama R, Maekawa M, Shimizu Y, Shimizu N, Liu JD, Wallace DC (1989) The human ATP synthase beta subunit gene: Sequence analysis, chromosomal assignment and differential expression Genomics 5: 829–843
Pinkel D, Landegent J, Collins C, Fuscoe J, Segraves R, Lucas J, Gray J (1988) Fluorescence in situ hybridisation with human chromosome-specific libraries: Detection of trisomy 21 and translocations of chromosome 4 Proc Natl Acad Sci USA 85: 9138–9142
Schmitz A, Olschwang S, Chaput B, Thomas G, Frelat G (1989) Oncogene detection by enzymatic amplification on flow sorted chromosomes Nucleic Acids Res 17: 816
Telenius H, Pelmear AH, Tunnacliffe A, Carter NP, Behmel A, Ferguson-Smith MA, Nordenskjold M, Pfragner R, Ponder BAJ (1992a) Cytogenetic analysis by chromosome painting using degenerate oligonucleotide- primed-polymerase chain reaction amplified flow-sorted chromosomes Genes, Chromosomes Cancer (in press)
Telenius H, Carter NP, Bebb CE, Nordenskjold M, Ponder BAJ, Tunnacliffe A (1992b) Degenerate oligonucleotide-primed PCR (DOP-PCR): general amplification of target DNA by a single degenerate primer Genomics (in press)
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1993 Springer-Verlag Berlin Heidelberg
About this paper
Cite this paper
Carter, N.P. (1993). Gene Mapping and PCR Applications with Flow-Sorted Chromosomes. In: Jacquemin-Sablon, A. (eds) Flow Cytometry. NATO ASI Series, vol 67. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-84616-8_22
Download citation
DOI: https://doi.org/10.1007/978-3-642-84616-8_22
Publisher Name: Springer, Berlin, Heidelberg
Print ISBN: 978-3-642-84618-2
Online ISBN: 978-3-642-84616-8
eBook Packages: Springer Book Archive