Topological Organization of Drosophila Hox Genes Using DNA Fluorescent In Situ Hybridization
DNA fluorescent in situ hybridization (FISH) is the method of choice to study genomic organization at the single-cell level. It has been recently used to study the topological organization of the homeotic bithorax complex (BX-C) in Drosophila as well as to describe long-range genomic interactions between the BX-C and the Antennapedia complex (ANT-C), in addition to other genomic loci. Coupled with immunofluorescence, FISH can be used to study the relative positioning of homeotic genes with nuclear subcompartments, such as Polycomb-group (PcG) bodies, transcription factories, or the nuclear lamina. Here, we describe two multicolor 3D-FISH protocols; one for whole mount Drosophila embryos or larval discs and one for Drosophila-cultured cells. Both methods can be applied to any single copy locus of interest and are compatible with immunostaining (FISH-I).
Key wordsFluorescent in situ hybridization Immunostaining Nuclear organization Gene positioning Chromosome topology Nuclear bodies Bithorax complex Antennapedia complex Polycomb-group proteins Drosophila
F.B. is supported by the CNRS. Research at the G.C. lab was supported by grants from the European Research Council (ERC-2008-AdG No 232947), the CNRS, the European Network of Excellence EpiGeneSys, the Agence Nationale de la Recherche and the Association pour la Recherche sur le Cancer.
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