GSM3518695: HCASMC SRF ChIPseq [TQ40]; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
SRF
Cell type
Cell type Class
Cardiovascular
Cell type
HCASMC
NA
NA
Attributes by original data submitter
Sample
source_name
Cultured coronary artery smooth muscle cells
tissue
Cultured HCASMC
growth medium
Grown in serum
chip antibody
anti-SRF (Santa Cruz sc-335)
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Briefly, approximately 4e6 HCASMC cells were fixed with 1 % formaldehyde and quenched by glycine. The cells were washed three times with PBS and then harvested in ChIP lysis buffer (50 mM Tris-HCl, pH 8, 5 mM EDTA, 0.5 % SDS). Crosslinked chromatin was sheared for 3x1 min by sonication (Branson SFX250 Sonifier) before extensive centrifugation. Four volumes of ChIP dilution buffer (20 mM Tris-HCl, pH 8.0, 150 mM NaCl, 2 mM EDTA, 1 % Triton X-100) was added to the supernatant. The resulting lysate was then incubated with Dynabeads™ Protein G (Thermo Scientific, 10009D) and antibodies at 4°C overnight. Beads were washed once with buffer 1 (20 mM Tris pH 8, 2 mM EDTA, 150 mM NaCl, 1% Triton X100, 0.1% SDS), once with buffer 2 (10 mM Tris pH 8, 1 mM EDTA, 500 mM NaCl, 1% Triton X100, 0.1% SDS), once with buffer 3 (10 mM Tris pH 8, 1 mM EDTA, 250 mM LiCl, 1% NP40, 1% sodium deoxycholate monohydrate) and twice with TE buffer. DNA was eluted by ChIP elution buffer (0.1 M NaHCO3, 1 % SDS, 20 μg/ml proteinase K). The elution was incubated at 65 °C over-night and DNA was extracted with DNA purification kit (Zymo D4013). The purified DNA was assayed by quantitative PCR with ABI ViiA 7 and Power SYBR Green Master Mix (ABI 4368706). Two biological replicates DNA was combined for library preparation. Libraries were prepared with KAPA Hyper Prep kit (KK8502).