GSM1827553: Lamin B1, Replicate 1; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
LMNB1
Cell type
Cell type Class
Lung
Cell type
IMR-90
Primary Tissue
Lung
Tissue Diagnosis
Normal
Attributes by original data submitter
Sample
source_name
IMR90 fibroblasts in culture
cell line
IMR90
antibody
Abcam #ab16048
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Cells were crosslinked with 1% formaldehyde diluted in PBS, without the addition of other co-crosslinkers, for 5 min at room temperature. After glycin quenching, the cell pellets were lysed in buffer containing 50 mM Tris, pH 7.5, 150 mM NaCl, 1% Triton, 0.1% Na-Deoxycholate, 0.1% SDS, supplemented with complete protease inhibitor cocktail (Thermo Scientific), and sonicated with Covaris sonicator, resulting in chromatin fragments of 250 bp average size. The supernatant was diluted 10 times with the above buffer without SDS, and subjected to immunoprecipitations with 2 ug of antibody or control IgG conjugated with Dynabeads Protein A or G (Invitrogen) at 4 °C for overnight. The beads were then washed 5 times with buffer containing 50 mM Tris, pH 7.5, 150 mM NaCl, 1% Triton, and 1 time with final wash buffer (50 mM Tris, pH 8.0, 10 mM EDTA, 50 mM NaCl), followed by elution with incubation of elution buffer (final wash buffer plus 1% SDS) at 65 °C for 30 min with agitation in a thermomixer. The ChIP and input were purified and used for qPCR analysis or for constructing sequencing libraries with the NEBNext Ultra kit (New England Biolabs).