Three-dimensional reconstruction of pericentromeric (1q12) DNA and ribosomal RNA sequences in HL60 cells after double-target in situ hybridization and confocal microscopy

Abstract

A fluorescent in situ hybridization procedure was applied to simultaneously label intranuclear pericentromeric (1q12) sequences of the chromosomes 1 and cytoplasmic ribosomal RNA sequences in whole cells of the promyelocytic HL60 cell line. For this purpose biotinated chromosome 1‐specific (1q12) repetitive satellite DNA and 28S ribosomal ssRNA probes were used. The entire procedure was performed in suspension to preserve nuclear morphology. The result was studied by three‐dimensional analysis, as provided by a scanning laser confocal microscope. The intracellular positions of both cytoplasmic rRNA and intranuclear centromere 1 DNA could easily be distinguished. This approach could be useful as a framework for the study of the 3‐D localization of genes and gene transcripts.