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Sample GSM1295076 Query DataSets for GSM1295076
Status Public on Dec 19, 2013
Title CBX6 BF ChipSeq (biological replicate 1 and 2)
Sample type SRA
Source name Cultured fibroblasts
Organism Homo sapiens
Characteristics tissue: normal breast
chip antibody: MRO19 - in house rabbit polyclonal
strain: BF
Treatment protocol Fibroblasts (8×105 cells per 10 cm dish) were infected with packaged LKO lentivirus vector containg CBX7shRNA (Sigma NM_175709.1-153s1c1). Infected cells were selected in puromycin (0.5ug/ml) and harvested after 10-14 days
Growth protocol The BF and Hs68 strains of human fibroblasts (30 population doublings) were cultured in Dulbecco modified Eagles medium (DMEM) supplemented with 10% fetal calf serum (FCS), 100 IU/ml penicillin, and 100 μ/ml streptomycin.
Extracted molecule genomic DNA
Extraction protocol Chip-Seq: DNA samples were end repaired, poly-A tailed and Illumina single end adapters were ligated following the standard Illumina protocol with minor adjustments. Agencourt AMPure XP beads at 0.8x ratio were used to size select out adapter dimers after adapter ligation. The Illumina kit Phusion enzyme was replaced by Kapa HiFi HotStart ready mix. Post PCR, AMPure XP beads were used at a 1:1 ratio to maintain size integrity and to allow use of the Invitrogen SizeSelect E-gel system. Samples were finally purified with QAIquick gel extraction kit and quality controlled on the DNA 1000 BioAnalyser 2100 chip before clustering and subsequent 36bp single end sequencing on the GAIIx analyser.
RNA-Seq: Total RNA was extracted using the High Pure RNA extraction Kit (Roche). RNA samples were quality controlled (QC) using the 6000 Nano RNA Chip on the BioAnalyser 2100 (Agilent) and subjected to poly-A selection using Sera-Mag oligo dT beads (Thermo Fisher Scientific Inc.). Libraries were prepared using the Directional mRNA-Seq Library Prep. v1.0. Pre-Release Protocol from Illumina with minor adjustments. The Illumina kit Phusion enzyme was replaced by Kapa HiFi HotStart ready mix which reduced the overall volume of the PCR and the ratio for the Agencourt AMPure XP beads was adjusted accordingly. The standard PCR cycling was also changed to match the concentration of the total RNA from the initial QC. After passing the final QC, the libraries were subjected to cluster formation and then 72bp single end sequencing on the GAIIx analyser.
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina Genome Analyzer IIx
Data processing ChipSeq: Duplicates were removed using the Picard MarkDuplicates program (picard-tools package version 1.48; with default parameters.
RNASeq: Alignments were performed to the UCSC version of the human transcriptome (Illumina iGenomes resource) using RSEM (version 1.2.4; Li and Dewey 2011).
Submission date Dec 19, 2013
Last update date May 15, 2019
Contact name Gordon Peters
Phone +44 207 269 3049
Organization name CRUK London Research Institute
Lab Molecular Oncology
Street address 44 Lincolns Inn Fields
City London
ZIP/Postal code WC2A 3LY
Country United Kingdom
Platform ID GPL10999
Series (1)
GSE40740 Genome wide co-localization of Polycomb orthologs and their effects on gene expression in human fibroblasts
BioSample SAMN02471437
SRA SRX396568

Supplementary file Size Download File type/resource
GSM1295076_CBX6_BF_ChipSeq_mergedReps_genome.wig.gz 623.7 Mb (ftp)(http) WIG
GSM1295076_CBX6_BF_ChipSeq_mergedReps_peaks.bed.gz 20.4 Kb (ftp)(http) BED
SRA Run SelectorHelp
Processed data provided as supplementary file
Raw data are available in SRA

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