title: COMBO-FISH for focussed fluorescence labelling of gene domains: 3D-analysis of the genome architecture of abl and bcr in human blood cells.
abstract: Structural analysis and nanosizing of gene domains requires not only high-resolution microscopy but also improved techniques of fluorescence labelling strongly focussed on the gene domains. To investigate the architecture of abl and bcr in blood cell nuclei forming the Philadelphia chromosome in CML, we applied COMBO-FISH using specifically colocalising combinations of triple strand forming oligonucleotide probes for abl on chromosome 9 and bcr on chromosome 22. Each probe set consisting of 31 homopyrimidine oligonucleotides was computer selected from the human genome database. Measurements by 3D microscopy were compared to results obtained after standard FISH using commercially available abl/bcr BAC probes. The relative radial fluorescence distributions in lymphocyte cell nuclei of healthy donors in comparison to cell nuclei of blood cells of CML patients showed a strong correlation in the location of abl and bcr for both labelling techniques. The absolute distances of the homologous bcr domains and the abl domain-nuclear center-abl domain angles in cell nuclei of CML donors differed significantly from those of healthy donors only when COMBO-FISH was applied. These results indicate that COMBO-FISH may be more sensitive than standard FISH in case of slight modifications in the genome architecture.
authors.affiliation: Kirchhoff-Institute of Physics, University of Heidelberg, Im Neuenheimer Feld 227, D-69120 Heidelberg, Germany.
authors.forename: Jutta
authors.initials: J
authors.lastname: Schwarz-Finsterle
authors.forename: Stefan
authors.initials: S
authors.lastname: Stein
authors.forename: Constance
authors.initials: C
authors.lastname: Grossmann
authors.forename: Eberhard
authors.initials: E
authors.lastname: Schmitt
authors.forename: Helmut
authors.initials: H
authors.lastname: Schneider
authors.forename: Luba
authors.initials: L
authors.lastname: Trakhtenbrot
authors.forename: Gideon
authors.initials: G
authors.lastname: Rechavi
authors.forename: Ninette
authors.initials: N
authors.lastname: Amariglio
authors.forename: Christoph
authors.initials: C
authors.lastname: Cremer
authors.forename: Michael
authors.initials: M
authors.lastname: Hausmann
mesh_terms: D020650:Combinatorial Chemistry Techniques / Q000379:methods*; D016044:Fusion Proteins, bcr-abl / Q000097:blood*; D016313:Genes, abl; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics* / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010677:Philadelphia Chromosome; D051562:Proto-Oncogene Proteins c-bcr / Q000097:blood / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods; D014178:Translocation, Genetic
chemical_list: D016044:Fusion Proteins, bcr-abl; C496613:BCR protein, human; D051562:Proto-Oncogene Proteins c-bcr