Sample information curated by ChIP-Atlas

Antigen

Antigen Class
Input control
Antigen
Input control

Cell type

Cell type Class
Lung
Cell type
NCI-H1299
Primary Tissue
Lung
Tissue Diagnosis
Carcinoma Non-Small Cell

Attributes by original data submitter

Sample

source_name
NCI-H1299
cell line
NCI-H1299
antibody
none
shRNA
NSD2_sh3

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
For ChIP-seq assays were performed according to the Millipore protocol. Cells were fixed using 1% formaldehyde, harvested, resuspended in ChIP lysis buffer (1% SDS, 10mM EDTA, 50mM Tris-HCl, pH 8.1) and sonicated using Bioruptor Pico (Diagenode) to generate fragments of 150 to 300 bp. Soluble chromatin was diluted 8 fold in ChIP RIPA buffer (10mMTris–HCl, pH 7.5, 140mMNaCl, 1mMEDTA, 0.5mMEGTA,1%Triton X-100, 0.1% SDS, 0.1% Na-deoxycholate) and incubated with Dynabeads Protein A (Invitrogen) coupled to H3K36me2 antibody ab9049 from Abcam. After incubation, the immunocomplexes were washed sequentially with Low Salt Wash Buffer (0.1% SDS, 1% Triton X-100, 2mM EDTA, 20mM Tris-HCl, pH 8.1, 150mM NaCl), High Salt Wash Buffer (0.1% SDS, 1% Triton X-100, 2mM EDTA, 20mM Tris-HCl, pH 8.1, 500mM NaCl), LiCl Wash Buffer (0.25M LiCl, 1% NP40, 1% deoxycholate, 1mM EDTA, 10mM Tris-HCl, pH 8.1) and TE. Immunocomplexes were eluted in ChIP elution buffer (1%SDS, 0.1M NaHCO3) and the crosslinking was reverted overnight at 65 ºC. Samples were treated with Proteinase K and RNase A and DNA was extracted using the QIAGEN PCR purification kit. Purified chromatin was used for library construction. For ChIP-seq the amount of DNA used was 15ng from each sample. Samples were processed through subsequent enzymatic treatments of end-repair, dA-tailing, and ligation to adapters with "NEXTflex ChIP Sequencing kit" from Bioo Scientific (part # 5143). Adapter-ligated libraries were completed by limited-cycle PCR (13 cycles) and extracted with a double double-sided SPRI size selection. Libraries were applied to an Illumina flow cell for cluster generation and sequenced on an Illumina instrument (see below) by following manufacturer's protocols.

Sequencing Platform

instrument_model
Illumina HiSeq 2000

hg38

Number of total reads
53585843
Reads aligned (%)
96.9
Duplicates removed (%)
7.1
Number of peaks
1244 (qval < 1E-05)

hg19

Number of total reads
53585843
Reads aligned (%)
96.0
Duplicates removed (%)
8.7
Number of peaks
1323 (qval < 1E-05)

Base call quality data from DBCLS SRA