Nevin Manimala Statistics

High-Throughput DNA FISH (hiFISH)

Methods Mol Biol. 2022;2532:245-274. doi: 10.1007/978-1-0716-2497-5_12.


High-throughput DNA fluorescence in situ hybridization (hiFISH) combines multicolor combinatorial DNA FISH staining with automated image acquisition and analysis to visualize and localize tens to hundreds of genomic loci in up to millions of cells. hiFISH can be used to measure physical distances between pairs of genomic loci, radial distances from genomic loci to the nuclear edge or center, and distances between genomic loci and nuclear structures defined by protein or RNA markers. The resulting large datasets of 3D spatial distances can be used to study cellular heterogeneity in genome architecture and the molecular mechanisms underlying this phenomenon in a variety of cellular systems. In this chapter we provide detailed protocols for hiFISH to measure distances between genomic loci, including all steps involved in DNA FISH probe design and preparation, cell culture, DNA FISH staining in 384-well imaging plates, automated image acquisition and analysis, and, finally, statistical analysis.

PMID:35867253 | DOI:10.1007/978-1-0716-2497-5_12

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