Chip seq high sensitivity kit

Human SH-SY5Y neuroblastoma cells were purchased from American Type Culture Collection (ATCC, Manassas, VA) and cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS) and 1% penicillin/streptomycin (Gibco-Invitrogen, Carlsbad, CA). For transient transfection, cells were transfected with indicated amounts of either flag-PARIS or flag using Fugene HD (Promega, Madison, WI) according to manufacturer’s instructions. After 48 h incubation, SH-SY5Y cells were cross-linked with 1% formaldehyde for 10 min at room temperature, and 125 mM glycine were then added to inactivate the formaldehyde. CHIP-seq high sensitivity kit (ab185908, Abcam) was further used for ChIP-Seq library preparation according to manufacturer’s instructions. Libraries were sequenced on an Illumina HiSeq 2000 sequencer.

A ChIP-Seq High Sensitivity Kit (Abcam, Cambridge, UK, ab185908) was prepared for this part. Generally, the HGC-27 cells were fixed by formaldehyde and lysed to extract the chromatin. After the chromatins were sheared, the RARα antibody was added to generate the immune complex. Next, the complexes were pelleted, and DNA was released and purified. Finally, qRT-PCR was performed for the input results. The sequence of primer was as follows: Site 3Forward: 5’-TAATAGGTGGTTTGAGGGC-3’
Reverse: 5’-TTCAGAAAGCATGCAGGTG-3’