Three-dimensional analysis of the organization of human chromosome domains in human and human-hamster hybrid interphase nuclei.

This report describes the intranuclear organization of chromosomes in human-hamster hybrid nuclei and in human cell nuclei. The target chromosomes were stained using in situ hybridization with biotinylated, chromosome-specific DNA probes. Bound probe was detected with fluorescein-avidin. Hybridizations were performed to fixed nuclei in aqueous suspension in order to preserve their three-dimensional morphology. Total nuclear DNA was stained with DAPI. Three-dimensional information about the organization of DNA and probe within the nucleus was obtained by optical sectioning. The human chromosomes in human-hamster hybrid nuclei were found to be confined to 'domains' that were maintained during the cell cycle. Different spatial localization patterns of the human chromosomes were seen in interphase nuclei of two different hybrid cell lines. The positions of chromosome-specific repetitive sequences in human fibroblast interphase nuclei were also studied using probes for the telomeric region of chromosome 1p (1p36), the centromeric region of chromosome 9 (9q12) and the long arm of the Y chromosome (Yq12). These studies showed that the two 1p telomeric loci are located near the nuclear surface. The chromosome 9 centromeric loci are similarly located. Simultaneous hybridization of the chromosome 1 telomeric probe (target size approximately 200 kb; b, base) and the Y-specific probe (target size greater than 2Mb), demonstrate that the binding sites of the two probes can be distinguished in the same nucleus on the basis of domain size.

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