GSM4271082: Calu3 RNAPII ChIP-seq Rep2; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
RNA polymerase
Antigen
RNA polymerase II
Cell type
Cell type Class
Lung
Cell type
Calu-3
Primary Tissue
Lung
Tissue Diagnosis
Adenocarcinoma
Attributes by original data submitter
Sample
source_name
Calu3
cell line
Calu3
cell type
Human lung adenocarcinoma cells
chip antibody
RNA polymerase II (RNAPII) Rpb1 NTD (D8L4Y) (Cell Signaling 14958S)
growth media
DMEM + 10% FBS
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Cells were lysed in lysis buffer (5mM PIPES pH 8.0, 85mM KCL, 0.5% NP-40, 1x Protease Inhibitor Cocktail (Roche)). Nuclei were isolated by centrifugation, lysed in RIPA buffer (1x PBS, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, Protease Inhibitor Cocktail), and sonicated. Protein-DNA complexes were isolated with antibodies. DNA ends were blunted using T4 DNA polymerase, Klenow DNA polymerase, and T4 polynucleotide kinase. Next, DNA was incubated with Klenow exo- to add 5' adenine overhangs. Multiplex adaptors were then ligated to the ends and converted to dsDNA using 5 cycles of PCR. DNA was size-selected to contain fragments between 200 and 300bp and PCR amplified for 5 cycles. DNA was purified with 1.2x AMPure XP beads. Sequencing was performed on an Illumina Hi-Seq.