title: GSM2417989: Mnase-seq.BG3C2.Ctrl1.fastq; Drosophila melanogaster; MNase-Seq
library_construction_protocol: Genomic DNA was prepared following regular protocol as described previously4 with some modifications. Add 600 μl of lysis buffer containing 100mM Tris-HCl, pH 8.5, 5mM EDTA pH8.0, 0.2% SDS, 200mM NaCl, and 20-25 μl of protease K @ 20mg/ml to the ground tissues or cells, mix well, and incubate at 55°C overnight. After the overnight digestion was cooled down to room temperature, add 5 μl of RNase A solution @ 20mg/ml for at least 1hr at RT. Following extraction with equal volume of phenol: chloroform: isoamyl alchohal =25:24:1, precipitation was carried out using equal volume of isopropanol. If flocky DNA forms, transfer it to a new tube containing 1ml of 70% ethanol. If necessary, spin down to collect the remained DNA in the precipitation solution followed by washing with 70% ethanol. If no flocky DNA appears, spin down @maximum speed for 10 min to collect gDNA pellet. 25ng of input genomic DNA or experimental enriched DNA were utilized for each library construction. 150-300 bp DNA fragments were selected by AMPure XP Beads (Beckman Coulter) after the adapter ligation. An Agilent 2100 BioAnalyzer was used to quantify the amplified DNA, qPCR was applied to accurately quantify the library concentration. 20 pM diluted libraries were used for sequencing. 50-cycle single-end sequencings were performed using Illumina HISeq 2000. Image processing and sequence extraction were done using the standard Illumina Pipeline. RNA-seq libraries were generated from duplicated samples per condition using the Illumina TruSeq RNA Sample Preparation Kit v2 following manufacturer’s protocol. The RNA-seq libraries were sequenced as 50-cycle pair-end runs using Illumina HiSeq 2000.
attributes.tag: GEO Accession
attributes.value: GSM2417989