GSM1969058: KDM5c Spikein Input Pooled CN rep2; Mus musculus; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
Input control
Antigen
Input control
Cell type
Cell type Class
Embryo
Cell type
Embryonic brains
NA
NA
Attributes by original data submitter
Sample
source_name
KDM5c_Spikein_Input_Pooled_CN
genotype/variation
Pooled Input (WT + SMCX KO)
gender
male
developmental stage
E16.5
tissue
mouse embryonic cortex
culture
10 Days in vitro
molecule subtype
Input-DNA
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
A small amount of sonicated drosphila chromatin was spiked-in to chromatin from mouse neurons. Spiked-In Chromatin was spiked-in with an antibody specific to drosphila specific histone variant. Chromatin was immunoprecipitated (overnight, 4°C) with beads and antibody (KDM5c 333). After immunoprecipitation, beads were washed as described previously. Immune complexes were eluted from beads (65°C, 5 min; and room temperature, 15 min) with 50 mM Tris-HCl pH 8.0, 1 mM EDTA and 1% SDS. Elution was repeated and eluates pooled. Reverse cross-linking was carried out (16h, 65°C) with addition of NaCl and RNase A. EDTA was increased to 5 mM and samples were incubated with 200 μg/ml proteinase K (2h, 50°C). DNA was recovered by phenol-chloroform extraction and ethanol precipitation, and quantifed by Qubit (Life Technologies). ChIP-Seq libraries were prepared through standard Illumina's procedure for library-preparation using NEBNext library reagents. ChIP-seq libraries were prepared from 10 ng DNA using NEBNext DNA library preparation reagents (E6000) and the protocol and reagent concentrations described in the Illumina Multiplex ChIP-seq DNA sample Prep Kit. Libraries were indexed using a single indexed PCR primer. After PCR amplification, 300-600 bp DNA fragments were selected on an agarose gel. Libraries were quantified by Qubit (Invitrogen), and library size was assessed by Bioanalyzer (Agilent). Libraries were sequenced using a HiSeq 2000 (Illumina) to generate 50 base single-end reads. ChIP-seq libraries were generated from two biological repeats per antibody, and matched Input samples.