GSM3629871: Snail1-HA ChIP-seq 1; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
Epitope tags
Cell type
Cell type Class
Digestive tract
Cell type
Colorectal adenocarcinoma
NA
NA
Attributes by original data submitter
Sample
source_name
colorectal adenocarcinoma cell line
cell type
pN1-LS174T Snail1-HA cells
chip antibody
abcam anti-HA (ChIP-grade) ab9110
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
For immunoprecipitation, aliquots of 200 µg crosslinked chromatin were used. Each aliquot was filled up to 200 µl with nuclei lysis buffer and then diluted with sonication buffer to a final volume of 1 ml. Magnetic Protein-G beads, preblocked for at least 4 h with 250 µg/ml BSA, and the appropriate antibody (listed in 4.10.1) were added to the diluted chromatin. The samples were incubated on a rotating platform at 4°C overnight. The next day, beads were collected by using a magnetic rack. Beads were washed two times with sonication buffer, two times with buffer A, two times with buffer B and two times with TE buffer. Each time, 1 ml of buffer was used for washing and the samples were incubated for 10 min on a rotating platform at 4°C. After the last washing step, bound material was eluted from the beads in two sequential rounds by adding 150 µl elution buffer and incubating each time at 67°C for 10 min with continuous shaking. The two eluates were combined, 10 µg RNAse and 90 µMol NaCl added and the mix was incubated for 5 h at 67°C with gentle agitation to reverse the crosslinks, followed by addition of 80 µg Proteinase K and incubation for 1 h at 45°C. For the input sample, used as reference material, 20% of the initial chromatin was filled up to 300 µl with elution buffer and processed like the immunoprecipitated sample. DNA was purified using the Qiagen PCR Purification Kit or PeqGold Cycle Pure Kit and stored at -20°C until needed. DNA libraries were prepared for 50 bp single-end sequencing on an Illumina HiSeq2000 system using standard Illumina protocols.