GSM6019612: Vitro_biotin; Oryza sativa; ChIP-Seq
The fragmented chromatin was diluted in G4DP buffer (10mM Tris-HCl, 5mM EDTA, 150mM KCl, 0.5mM DTT and 0.5% NP-40, pH=7.4) until the final concentration of SDS was below 0.1%. 1/10 volume of supernatant was kept as input. 30 μl of 1 mM BioTASQ was added into diluted chromatin and incubated overnight at 4 °C, then 30 μl of washed M280 Streptavidin Dynalbeads (11206D, Invitrogen) was added and incubated for another 4 h at 4°C. The washed BioTASQ/BioCyTASQ-bound DNA was eluted with 200 μl elution buffer at 65°C for two times with 15 min each, followed by de-crosslinking at 65°C overnight. De-crosslinked G4DPed DNA was extracted using phenol/chloroform followed by cold ethanol precipitation. 30 nm biotin (Sigma, cat, no. 2031) was used as control. Two biological replicates were conducted for BioTASQ/BioCyTASQ and biotin only. All G4DPed DNA was used for the library preparation, which was constructed using the NEBNext® Ultra™ II DNA Library Prep Kit for Illumina (NEB, E7645S). All libraries were sequenced using paired-end mode on Illumina NovaSeq platform