GSM1815465: GAFm_MNase; Drosophila melanogaster; MNase-Seq
MNase-digested chromatin was prepared as described previously (Petesch and Lis 2008; Gilchrist et al. 2009; Gilchrist et al. 2010) except that 200 µl chromatin was digested with 20 units MNase (Worthington) for 3 hours at 25 degrees C. Following gel purification, mono-nucleosome sized fragments (100-200 bp) were subjected to paired-end sequencing using the Illumina paired-end protocol. ChIP-seq libraries were prepared using the Illumina ChIP-seq Library preparation Kit following the manufacture’s instructions with minor modifications. Briefly 10ng of purified ChIP DNA was end repaired by conversion of overhangs to phosphorylated blunt ends. A’ bases were added to the 3’ends of the DNA fragments and Illumina adapters (1:30 dilution) were ligated to the ends of ChIP fragments.  This fragment range corresponds to a ChIP fragment range of about 200 -300bp. Size selected fragments were PCR amplified for 12 cycles . Libraries were quantified and validated using Agilent Technologies 2100 Bioanalyser for size, concentration and purity. Q-PCR was repeated to confirm retention of relative enrichment.