title: GSM1402302: Batch1_ChIP-seq_PGAL::TOP1rnh1drnh201d; Saccharomyces cerevisiae; ChIP-Seq
library_construction_protocol: Sonicated and formaldehyde-crosslinked-chromatin were treated with antibody S9.6 for immunoprecipitation of RNA-DNA hybrids (ChIP). Libraries were prepared according to Illumina's instructions with modifications. Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 18 cycles and library fragments of ~250-350 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced using Illumina MiSeq Benchtop Sequencer following the manufacturer's protocols. S9.6 CHIP-seq and input chromatin (=control sonicated-crosslinked-DNA)
identifiers.namespace: GEO
identifiers.id: GSM1402302