GSM1542534: Input NCI-H2052 rep1; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
Input control
Antigen
Input control
Cell type
Cell type Class
Lung
Cell type
H2052
Primary Tissue
Lung
Tissue Diagnosis
Adenocarcinoma
Attributes by original data submitter
Sample
source_name
Mesothelioma cells
cell line
NCI-H2052
cell type
malignant pleural mesothelioma cells
chip antibody
none
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Cells were cross-linked in the presence of 1% formaldehyde for 10 min at at room temperature. Crosslinking was stopped by addition of 125mM Glycine for 5 min. Cells were rinsed with 1x PBS and scraped off. Pellets were resuspended in buffer 1 (10 mM Tris (pH 8.0), 10 mM EDTA, 0.5 mM EGTA, 0.25% Triton X-100) and washed once buffer 2 (10 mM Tris (pH 8.0), 1 mM EDTA, 0.5 mM EGTA, 200 mM NaCl). Then cells were lysed in lysis buffer (50 mM HEPES/KOH (pH 7.5), 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% DOC, 0.1% SDS, protease inhibitors) and sonicated for 15min using a Covaris E210 (TR24-M (500035) and TR12x12 Tubes (520080), Settings: 5% duty cycle, intensity 4). 70ug of chromatin were incubated over night at 4°C with 7ul YAP antibody (Abcam, [EP1674Y] (ab52771) or 20ul TEAD1 antibody (BD Transduction LaboratoriesT (610922)), and for 2h with 25ul preblocked (tRNA, BSA) Dynabeads protein G. Beads were washed twice with lysis buffer, once with DOC buffer (10 mM Tris (pH 8.0), 0.25 M LiCl, 0.5% NP-40, 0.5% deoxycholate, 1 mM EDTA) and once with TE pH8.0, and bound chromatin was eluted in 1% SDS/0.1 M NaHCO3. After RNase A treatment, cross-linking was reversed by overnight incubation at 65 °C followed by proteinase K digestion. DNA was purified using the Minielute PCR purification kit (Qiagen). A sample of the input chromatin was treated in the same way to generate total input DNA. Input and ChIP-samples in duplicate were sequenced. Libraries were prepared using the NEBNext ChIP-seq library reagent set for Illumina (New England BioLabs Inc.) omitting the size selection step before PCR enrichment.