title: GSM4290968: 9311-HS-1 MeDIP-Seq; Oryza sativa; MeDIP-Seq
library_construction_protocol: Genomic DNA was extracted from3-week-old wild-type Oryza sativa 93-11and sonicated to produce fragments of ~250 bp. Fragmented DNA was incubated for 2 h at 4 °C with anti-6mA polyclonal antibody (Synaptic Systems) in immunoprecipitation buffer (10 mM sodium phosphate, 2 mM potassium phosphate, 137 mM NaCl, 2.7 mM KCl, 0.05% Triton X-100, pH 7.4). The mixture was then immunoprecipitated by incubation with Protein A/G Plus-Agarose (Santa Cruz) that was pre-bound with bovine serum albumin (BSA) at 4 °C for additional 2 h. After extensive washing, bound DNA was eluted from the beads in elution buffer (50 mM Tris, pH 8.0, 1% SDS, 10 mM EDTA) at 65 °C, treated with proteinase K and purified using QIAquick PCR Purification Kit (Qiagen). DNA quality was monitored on agarose gels and Qubit® DNA Assay in Qubit® 2.0Flurometer (Life Technologies). Two biological replicates for 6mA IP and one biological replicates for input were prepared with NEBNext DNA library prep kit (NEB) following manufacturer's instructions, Size selection and gel purification, followed by PCR amplification. Library concentration was quantified by Qubit®2.0 Flurometer (Life Technologies) and the insert size was determined by Agilent Bioanalyzer 2100 system (Agilent).
attributes.tag: GEO Accession
attributes.value: GSM4290968