Computational protocol: NCoR controls glioblastoma tumor cell characteristics

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Protocol publication

[…] Chromatin immunoprecipitation (ChIP) was performed following the High Cell ChIP kit # protocol from Diagenode. Five micrograms of anti-NCoR from Abcam (ab24552) were used in each IP. ChIP analysis was done with Q-PCR using Invitrogen Platinum SYBR Green qPCR Supermix-UDG together with site-specific primers. For genome-wide sequencing ChIP sequencing (ChIP-seq) analysis, 5 μg of chromatin was used in 2 separate IP's and combined into one elution. Subsequently, the DNA sequencing library was made using a kit from Illumina, except Illumina TruSeq adaptors were used to enable multiplexing. The library was analyzed by Solexa/Illumina Hi-seq. After prefiltering the raw data by removing sequenced adapters and low-quality reads, the sequence tags were aligned to the human genome (assembly hg19) with a Bowtie alignment tool. To avoid any PCR-generated spikes, we allowed only one read per chromosomal position and thus eliminated PCR bias. From the filtered raw data, 2 million unique reads per sample were used for peak detection. Peak detection was performed using the CisGenome program with a 2-sample analysis where sequenced input (1%) was used as a negative control. Peaks were called with a window statistic cutoff of 3 and a log2 fold change of 2. […]

Pipeline specifications

Software tools Bowtie, CisGenome
Application ChIP-seq analysis
Organisms Homo sapiens
Diseases Glioblastoma, Neoplasms