GSM3020136: CHIPseq-MCF10A Epithelial-treated with Nutlin3A-CHIP for p53; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
TFs and others
Cell type Class
Attributes by original data submitter
Breast mammary gland
Sequenced DNA Library
For RNA-Seq: Total mRNA was extracted with E.Z.N.A. Total RNA kit (Omega; R6834-02) and Poly(A)+ RNA was isolated by double selection with poly-dT beads, using 2.5 mg total RNA, which is then followed by first- and second-strand synthesis For RNA-Seq: Libraries were prepared using NEXTflex Rapid Illumina DNA-Seq Library Prep Kit (Bioo Scientific). For ChIP-Seq: Cells were crosslinked at 80% confluency with 1% formaldehyde for 10 min at room temperature. Crosslinking was quenched with 125mM glycine and the resulting pellet was washed twice with cold PBS and lysed. Samples were subjected to sonication with Diagenode Bioruptor Plus for 40 cycles (30 sec on/off at high setting) for shearing chromatin to 150-500 bp average size. Immunoprecipitation reactions were performed with Diagenode IP-Star Compact Atuomated System with mentioned antibodies. Immunoprecipitated DNA was reverse-crosslinked at 65°C for 4 hours and then eluted from beads, purified using SPRI beads, and used to construct sequencing libraries. For ChIP-Seq: Libraries were prepared using NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs).