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Karyotyping human chromosomes by combinatorial multi-fluor FISH

Abstract

We have developed epifluorescence filter sets and computer software for the detection and discrimination of 27 different DNA probes hybridized simultaneously. For karyotype analysis, a pool of human chromosome painting probes, each labelled with a different fluor combination, was hybridized to metaphase chromosomes prepared from normal cells, clinical specimens, and neoplastic cell lines. Both simple and complex chromosomal rearrangements could be detected rapidly and unequivocally; many of the more complex chromosomal abnormalities could not be delineated by conventional cytogenetic banding techniques. Our data suggest that multiplex-fluorescence in situ hybridization (M-FISH) could have wide clinical utility and complement standard cytogenetics, particularly for the characterization of complex karyotypes.

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References

  1. Gray, J.W. et al. Molecular cytogenetics: diagnosis and prognostic assessment. Curr. Opin. Biotech. 3, 623–631 (1992).

    Article  CAS  PubMed  Google Scholar 

  2. Xing, Y. & Lawrence, J.B. Molecular cytogenetics: applications of FISH to chromosomal aberrations in cancer genetics. In The causes and consequences of chromosomal aberrations, (ed. Kirsch, I.R.) 3–28 (CRC Press, BocaRaton, 1993).

    Google Scholar 

  3. Nederlof, P.M. et al. Three color fluorescence in situ hybridization for the simultaneous detection of multiple nucleic acid sequences. Cytometry 10, 20–27 (1989).

    Article  CAS  PubMed  Google Scholar 

  4. Nederlof, P.M. et al. Multiple fluorescence in situ hybridization. Cytometry 11, 126–131 (1990).

    Article  CAS  PubMed  Google Scholar 

  5. Ried, T., Baldini, A., Rand, T.C. & Ward, D.C. Simultaneous visualization of seven different DNA probes by in situ hybridization using combinatorial fluorescence and digital imaging microscopy. Proc. Natl. Acad. Sci. USA 89, 1388–1392 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Dauwerse, J.G., Wiegant, J., Raap, A.K., Breuning, M.H. & van Ommen, G.J.B. Multiple colors by fluorescence in situ hybridization using ratio-labelled DNA probes create a molecular karyotype. Hum. Mol. Genet. 1, 593–598 (1992).

    Article  CAS  PubMed  Google Scholar 

  7. Nederlof, P.M., van der Flier, S., Vrolijk, J., Tanke, H.J. & Raap, A.K. Fluorescence ratio measurements of double-labeled probes for multiple in situ hybridization by digital imaging microscopy. Cytometry 13, 839–845 (1992).

    Article  CAS  PubMed  Google Scholar 

  8. du Manoir, S. et al. Detection of complete and partial chromosome gains and losses by comparative genomic in situ hybridization. Hum. Genet. 90, 590–610 (1993).

    Article  CAS  PubMed  Google Scholar 

  9. Ried, T., Landes, G., Dackowski, W., Klinger, K. & Ward, D.C. Multicolor fluorescence in situ hybridization for the simultaneous detection of probe sets for chromosomes 13, 18, 21, X and Y in uncultured amniotic fluid cells. Hum. Mol. Genet. 1, 307–313 (1992).

    Article  CAS  PubMed  Google Scholar 

  10. Lengauer, C. et al. Chromosomal bar codes constructed by fluorescence in situ hybridization with Alu-PCR products of multiple YAC clones. Hum. Mol. Genet. 2, 505–512 (1993).

    Article  CAS  PubMed  Google Scholar 

  11. Popp, S. et al. A strategy for the characterization of minute chromosome rearrangements using multiple color fluorescence in situ hybridization with chromosome specific DNA libraries and YAC clones. Hum. Genet. 92, 527–532 (1993).

    Article  CAS  PubMed  Google Scholar 

  12. Wiegant, J. et al. Multiple and sensitive fluorescence in situ hybridization with rhodamine-, fluorescein-, and coumarin-labeled DNAs. Cytogenet. Cell Genet. 63, 73–76 (1993).

    Article  CAS  PubMed  Google Scholar 

  13. Ernst, L.A., Gupta, R.K., Mujumdar, R.B. & Waggoner, A.S. Cyanine dye labeling reagents for sulfhydryl groups. Cytometry 10, 3–10 (1989).

    Article  CAS  PubMed  Google Scholar 

  14. Mujumdar, R.B., Ernst, L.A., Mujumbar, S.R. & Waggoner, A.S. Cyanine dye labeling reagents containing isothiocyanate groups. Cytometry 10, 11–19 (1989).

    Article  CAS  PubMed  Google Scholar 

  15. Yu, H. et al. Cyanine dye dUTP analogs for enzymatic labeling of DNA probes. Nucl. Acids Res. 22, 3226–3232 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Waggoner, A. Covalent labeling of proteins and nucleic acids with fluorophores. Meth. Enzym. 246, 362–373 (1995).

    Article  CAS  PubMed  Google Scholar 

  17. Francke, U. Digitized and differentially shaded human chromosome ideograms for genomic applications. Cytogenet. Cell Genet. 65, 205–219 (1994).

    Article  Google Scholar 

  18. Bray-Ward, P. et al. Integration of the cytogenetic, genetic and physical maps of the human genome by FISH mapping of CEPH YAC clones. Genomics (in the press).

  19. Baldini, A. & Ward, D.C. In situ hybridzation banding of human chromosomes with Alu-PCR products: a simultaneous karyotype for gene mapping studies. Genomics 9, 770–774 (1991).

    Article  CAS  PubMed  Google Scholar 

  20. Matera, A.G. & Ward, D.C. Oligonucleotide probes for the analysis of specific repetitive DNA sequences by fluorescence in situ hybridization. Hum. Mol. Genet. 1, 535–539 (1992).

    Article  CAS  PubMed  Google Scholar 

  21. Chumakov, I.M. et al. A YAC contig of the human genome. Nature 377, 175–299 (1995).

    CAS  PubMed  Google Scholar 

  22. Hudson, T.J. et al. An STS-based map of the human genome. Science 270, 1945–1954 (1995).

    Article  CAS  PubMed  Google Scholar 

  23. Meltzer, P.S., Guan, X.Y., Burgess, A. & Trent, J. Rapid generation of region specific probes by chromosome microdissection and their application. Nature Genet. 1, 24–28 (1992).

    Article  CAS  PubMed  Google Scholar 

  24. Guan, X.Y., Trent, J.M. & Meltzer, P.S. Generation of band-specific painting probes from a single microdissected chromosome. Hum. Mol. Genet. 2, 1117–1121 (1993).

    Article  CAS  PubMed  Google Scholar 

  25. Guan, X.Y., Meltzer, P.S. & Trent, J.M. Rapid generation of whole chromosome painting probes (WCPs) by chromosome microdissection. Genomics 22, 101–107 (1994).

    Article  CAS  PubMed  Google Scholar 

  26. Guan, X.Y., Meltzer, P.S., Burgess, A. & Trent, J.M. Complete coverage of chromosome 6 by chromosome microdissection: Generation of 14 band region-specific probes. Hum. Genet. 95, 637–640 (1995).

    Article  CAS  PubMed  Google Scholar 

  27. Guan, X.Y. et al. Chromosome arm painting probes. Nature Genet. 12, 10–11 (1996).

    Article  CAS  PubMed  Google Scholar 

  28. Ledbetter, D.H. The ‘colorizing’ of cytogenetics: is it ready for prime time? Hum. Mol. Genet. 5, 297–299 (1992).

    Article  Google Scholar 

  29. Waggoner, A. et al. Multiple spectral parameter imaging. Meth. Cell. Biol. 30, 449–478 (1989).

    Article  CAS  Google Scholar 

  30. du Manoir, S. et al. Quantitative analysis of .comparative genomic hybridization. Cytometry 9, 21–49 (1995).

    Google Scholar 

  31. Smith, T.G. Jr., Marks, W.B., Lange, G.D., Sheriff, W.H. & Neale, E.A. Edge detection in images using Marr-Hildreth filtering techniques. J. Neurosci. Meth. 26, 75–81 (1988).

    Article  Google Scholar 

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Speicher, M., Ballard, S. & Ward, D. Karyotyping human chromosomes by combinatorial multi-fluor FISH. Nat Genet 12, 368–375 (1996). https://doi.org/10.1038/ng0496-368

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