GSM4295300: w1118 FLAG IP control 1; Drosophila melanogaster; ChIP-Seq
ChIP-seq was performed on six-day-old w1118 controls and w1118; dERR-3XFLAG-BACVK22; dERR1/dERR2 adults essentially as described (Barry and Thummel 2016) using FLAG antibodies (Sigma F1804, M2 monoclonal). ChIP-seq library construction is performed using the Illumina TruSeq ChIP Sample Prep Kit (IP-202-1012 and IP-202-1024) as described below.  Briefly, ChIP DNA is quantitated using an Invitrogen Qubit dsDNA HS Assay (Q32851) and a quantity representing appoximately 10 ng is converted to blunt-ended fragments with 5'-phosphates and 3'-hydroxyl groups using a combination of enzymes that perform fill-in reactions and exhibit exonuclease activity.  An A-base is added to the blunt-ended DNA as a means to prepare the fragments for adapter ligation and to block concatamer formation during the ligation step.  Adapters containing a T-base overhang are ligated to the A-tailed DNA fragments.  Ligated fragments are PCR-amplified (10 cycles) and the amplified library is purified  using Agencourt AMPure XP beads.  The concentration of the amplified library is measured using the Invitrogen Qubit dsDNA HS Assay and an aliquot of the library is resolved on an Agilent 2200 Tape Station using a D1K (cat# 5067-5361 and 5067-5362) or a High Sensitivity D1K (cat# 5067-5363 and 5067-5364) assay to define the size distribution of the sequencing library.  Libraries are adjusted to a concentration of approximately 10 nM and quantitative PCR is performed using the KapaBiosystems Kapa Library Quant Kit (cat# KK4824) to calculate the molarity of adapter ligated library molecules.  The concentration is further adjusted following qPCR to prepare the library for Illumina sequence analysis.
GEO Accession
GSM4295300