GSM2449945: SH-SY-IgG ChIPseq in SHSY5Y; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
Input control
Antigen
Input control
Cell type
Cell type Class
Neural
Cell type
SH-SY5Y
Primary Tissue
Brain
Tissue Diagnosis
Neuroblastoma
Attributes by original data submitter
Sample
source_name
SHSY5Y
cell line
SHSY5Y
cell type
Midbrain Dopamine Neurons
chip antibody
IgG Sigma I5381
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
ChIP experiment was performed as previously reported with few modifications (Langlais et al, Molecular Cell 2012). Briefly, human SHSY5Y cells stably expressing mouse Pitx3-3Fn were grown to 90% confluence, fixed in 1% formaldehyde/PBS for 5min at room temperature, washed, scraped off from the petri and sonicated to yield chromatin fragments with average size of 300-600bp. Sheared chromatin was incubated overnight at 4˚C along with 1:1 mix of Dynabeads-Protein A:Dynabeads-Protein G (Invitrogen) already coupled to primary antibody and thoroughly washed. Primary antibodies that were used were against the native form of Pitx3 (rabbit polyclonal, home-made) and Pitx3-Flag forms (mouse FlagM2, Sigma), including also rabbit (G2018, Sigma) or mouse (I5381, Sigma) IgG as controls for specific antibodies. Immunoprecipitated chromatin was washed, de-crosslinked, and then briefly treated with proteinase K and RNAse A. DNA from input, specific antibody and IgG was then purified using Qiaquick PCR purification kit (Qiagen). Immunoprecipitated DNA was sent for sequencing on Illumina GAIIx or Hi-Seq 2000 according to McGill University and Genome Quebec Innovation Centre protocols.