title: GSM1303938: TOP1α_1; Arabidopsis thaliana; MNase-Seq
library_construction_protocol: 1g of inflorescences containing unopened buds from Ler and top1a-2 plants were ground in liquid nitrogen, and the powder was transferred to M1 buffer (10mM phosphate buffer pH7.0, 0.1M NaCl, 10mM mercaptoethanol, 1M hexylene glycol, 1x protease inhibitor cocktail (Roche) and 1mM PMSF). The suspension was filtered through four layers of miracloth. The filtrate was centrifuged at 13,400g for 10 min at 4°C. The pellet (crude nuclei preparation) was resuspended and washed in M2 buffer (M1 buffer plus 10 mM MgCl2 and 0.5%TritonX-100) three times and once in M3 buffer (10mM phosphate buffer pH7.0, 0.1M NaCl, 10mM mercaptoethanol, 1x protease inhibitor cocktail (Roche) and 1mM PMSF). After centrifugation at 16,400g for 1min at 4°C, the nuclei were dissolved in digestion buffer (50mMTris-HCl, 5mM CaCl2, 1xBSA, 20mg/ml RNase A, 1x protease inhibitor cocktail (Roche)) containing 0.4U/ul Micrococcal Nuclease (NEB) and incubated at 37°C for 30min. DNA was extracted using the CTAB method and resolved in a 2% agarose gel. DNA bands corresponding to ~150bp were cut and purified with the Gel Purification Kit (Qiagen). DNA bands corresponding to ~150bp were cut and purified with the Gel Purification Kit (Qiagen). DNA libraries were constructed with the TruSeq DNA Sample Preparation v2 kit (Illumina) and sequenced with Illumina's HiSeq2000 platform
attributes.tag: GEO Accession
attributes.value: GSM1303938