title: GSM3100198: HeLa WT, RNA-seq, rep1; Homo sapiens; RNA-Seq
library_construction_protocol: For human ChIP-seq, ~15 million wild-type HeLa S3 cells or the indicated hNDF knockout cells were collected and crosslinked with 1% formaldehyde. After cell lysis and genome fragmentation by sonication, samples were incubated with 20 ?L of Dynabeads Protein A that were pre-bound with 1 ?L of NDF antiserum. [Note that 1 ?L of hNDF antiserum was optimal for hNDF antiserum #7. For hNDF antiserum #6, 2 ?L was the optimal volume.] Bound DNA were isolated with MinElute columns (Qiagen). For human RNA-seq, total RNA was purified by using the PureLink RNA Mini RNA Kit (Thermo Fisher Invitrogen). Mouse BMDM ChIP-seq and RNA-seq were performed as described in Kaikkonen, M. U. et al. Remodeling of the enhancer landscape during macrophage activation is coupled to enhancer transcription. Mol. Cell 51, 310-325. Human ChIP-seq libraries were prepared by end-repair, A-tailing, and ligation of Illumina TruSeq adaptors, which was then followed by PCR amplification and size selection with AMPure beads (Beckman Coulter). Human RNA-seq libraries were prepared with the TruSeq Stranded mRNA Library Prep Kit (Illumina). Mouse ChIP-seq libraries were prepared from ChIP DNA and inputs by blunting, A-tailing, and adapter ligations were performed by using NextFlex barcode adapters. Mouse RNA-seq libraries were prepared from poly(A)-enriched mRNA, as described in Kaikkonen, M. U. et al. Mol. Cell 51, 310-325. GRO-seq libraries were prepared using NEBNext multiplex small RNA library prep set for Illumina.
attributes.tag: GEO Accession
attributes.value: GSM3100198