Chip it control kit human

HepG2 cells transfected with pCMVMycPod-1 were fixed with 1% formaldehyde for 10 min. ChIP assays were performed using the ChIP-IT Express kit (Active Motif, Rixensart, Belgium) following the manufacturer's instructions. Chromatin was fragmented by sonication with eight 10-s pulses at 25 μm amplitude in a VCX130PB ultrasonic processor (Sonics & Materials, CT, USA). Most resulting chromatin fragments ranged from 200 bp to 600 bp. Sheared chromatin was incubated with 3 μg anti-MYC (Clontech) or with 1 μg IgG as negative control (ChIP-IT Control Kit-Human, Active Motif).

Chromatin immunoprecipitation (ChIP) was performed applying ChIP-IT^® Express Enzymatic Kit (Active Motif, Carlsbad, CA, USA) precisely according to the manufacturer’s instructions. Briefly, cells were fixed with 1% formaldehyde, and chromatin was sheared by enzymatic shearing (10 min, 37 °C). Then, immunoprecipitation procedure was performed using 5 μL of anti-ChREBP antibody (cat. no ab92809, RRID:AB_10562135) per 10 μg of chromatin. ChIP-IT^® Control Kit—Human (Active Motif, Carlsbad, CA, USA) was used as a quality control. Both mRNA purification and PCR analysis were performed as described in Section 4.7.