GSM3465940: met1_input_dnase_rep3; Arabidopsis thaliana; DNase-Hypersensitivity
Arabidopsis nuclei were isolated and re-suspended in NIB as described in ChIP-seq library preparation. After centrifugation at 6,000 rpm for 20 min at 4 °C, nuclei pellets were re-suspended in 400 ml DNaseI buffer (10 mM Tris-HCl pH 8.0, 2.5 mM MgCl2, 0.5 mM CaCl2). 100 ml of nuclei was incubated with 0.1-2 U / ml DNase I (New England Biolabs, cat. No. M03030S) for 3 min at 37 °C. 50 mM EDTA, 0.5 % SDS (final concentration), 2 ml proteinase K were added and incubated at 55 °C for 1 hr to stop digestion. To completely denature chromatin, more SDS were added to make final 1% concentration then incubated at 65 °C for 15 min. gDNA were isolated with P:C:I and DNA fragmentation were visualized by DNA electrophoresis. Samples with similar degree of DNA digestion were selected to construct library. 0.5 X volume of SPRI beads were added to remove undigested gDNA. Supernatant were collected and purified with 1.25 X volume of SPRI beads.
GEO Accession
GSM3465940