GSM6943938: ChIP-RPB1-pS5_dTAG-3h_INTS8-dTAG_INTS8-N211del_DLD1_rep2; Homo sapiens; ChIP-Seq
Lysates were clarified from sonicated cells and RBP1-DNA complexes were isolated with antibody. Libraries were prepared according to Vazyme's instructions accompanying the VAHTS Universal DNA Library Prep Kit (Vazyme ND607-02). 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 (32 to 52 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 15 cycles and library fragments of ~250 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 on the Illumina NovaSeq 6000 following the manufacturer's protocols.