anti RNAPII-S7p 4E12 (Chapman et al. science 2007)
source
Fixed chromatin
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
ChIP for RNAPII modifications was performed using chromatin from ESCs fixed in 1% formaldehyde (Sigma) at 37°C for 10 min, as described (associated paper, Brookes et al., 2012; Stock et al., 2007). 600-700 μg of chromatin were used for immunoprecipitation. ChIP-seq libraries were prepared from 10 ng of immunoprecipitated genomic DNA (quantified by Picogreen and Qubit) using the Next ChIP-Seq Library Prep Master Mix Set from Illumina (NEB, # E6240) following NEB protocol with some modifications. Samples were PCR amplified prior to size selection on an agarose gel. Libraries were quantified by Qubit (Invitrogen) and qPCR Kapa Library Quantification Universal Kit (KapaBiosytems, # KK4824), and library size was assessed by Bioanalyzer (Agilent). mRNA libraries were made using TruSeq RNA Sample Preparation Kits v2 setA (Illumina, # RS-122-2001) following the manufacturer's instructions. Libraries were quantified by Qubit (Invitrogen) and qPCR Kapa Library Quantification Universal Kit (KapaBiosytems, # KK4824), and library size was assessed by Bioanalyzer (Agilent)