Quantitative localization of chromosomal loci by immunofluorescence

Abstract

DNA within the yeast nucleus is spatially organized. Yeast telomeres cluster together at the nuclear periphery, centromeres cluster together near the spindle pole body, and both the rDNA repeats and tRNA genes cluster within the nucleolus. Furthermore, the localization of individual genes to subnuclear compartments can change with changes in transcriptional status. As such, yeast researchers interested in understanding nuclear events may need to determine the subnuclear localization of parts of the genome. This chapter describes a straightforward quantitative approach using immunofluorescence and confocal microscopy to localize chromosomal loci with respect to well characterized nuclear landmarks.