GSM5280134: MN-SC88_S15; Saccharomyces cerevisiae; MNase-Seq
Mononucleosomal DNA was isolated as described in Gonzalez et al. (2016). Cells were fixed with 1% formaldehyde and treated with 10 mg of Zymolyase 20T for 5 minutes at 30º C to generate spheroplasts. These were resuspended in NP-buffer (1 M Sorbitol; 50 mM NaCl; 10 mM Tris pH7.4; 5 mM MgCl2; 1 mM CaCl2; 0.075% NP-40; 1 mM β-Mercaptoethanol; 500 uM Spermidine) and digested with 300 units/mL of micrococcal nuclease (MNase) at 37 ºC during 10 minutes. The amount of MNase was optimized experimentally for each strain to generate a 80:20 ratio of mononucleosome to dinucleosome fragments. Libraries of mononucleosomal DNA were constructed following the Illumina protocol and were sequenced in an Illumina NextSeq500 platform using the paired-end protocol. We generated between 13 and 32 million reads of 75 nt per experiment, representing a 83- to 203-fold genome coverage.