Normal rabbit IgG (Santa Cruz Biotechnology, sc-2027)
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Lats1/2-deficient (Lats1fl/fl;Lats2fl/+;CcGFPfl/+;Dhh-Cre) mouse paraspinal tumors were isolated, minced into 1 mm3 pieces and dissociated using collagenase type I (156 U/ml) and dispase protease II (1.25 U/ml) at 37oC. Tumor cells were propagated as an adherent layer on tissue culture dishes in proliferation medium composed of DMEM/10% FBS plus L-glutamine and penicillin/streptomycin. Dissociated Lats1/2-deficient mouse tumor cells (∼20 million cells) were fixed for 15 min at room temperature with 1% formaldehyde-containing medium. Nuclei were isolated and sonicated in sonication buffer (10 mM Tris-HCl [pH 8.0], 1 mM EDTA, 0.5 mM EGTA, and protease inhibitor cocktail). Sonicated chromatin (∼300 μg) was used for immunoprecipitation by incubation with rabbit IgG or anti-Tead1, anti-H3K27ac, anti-H3K4me3, anti-H3K4me1 antibody (4 μg) overnight at 4°C. The ChIP-seq libraries were prepared using NEBNext ChIP-seq Library Prep Master Mix Set for Illumina (NEB catalogue number E6240L) and then run on the Illumina sequencer HiSeq 2000.