Genome-wide mapping of nucleosome positions in yeast using high-resolution MNase ChIP-Seq

Abstract

Eukaryotic DNA is packaged into chromatin where nucleosomes form the basic building unit. Knowing the precise positions of nucleosomes is important because they determine the accessibility of underlying regulatory DNA sequences. Here we describe a detailed method to map on a genomic scale the locations of nucleosomes with very high resolution. Micrococcal nuclease (MNase) digestion followed by chromatin immunoprecipitation and facilitated library construction for deep sequencing provides a simple and accurate map of nucleosome positions.

Figure 10.1

MNase digestion results. The input lane indicates undigested chromatin prior to MNase treatment. (A) An MNase ladder is visible in all three MNase-treated supernatant lanes. As indicated in the gel, with increasing MNase concentration, there is an increase in mononucleosomal DNA and decrease in higher molecular weight bands corresponding to “polynucleosomes.” Left-most lane indicates a 1-kb ladder. (B) The pellet lane indicates a small amount of unsolubilized DNA trapped in the pellet prior to sonication.

Figure 10.2

Agarose gel purification. Regions A and C indicate 1-kb ladder. Region B: red box indicates the excised gel portion.