alias: GSM4705423
title: GSM4705423: idn2_r2_ATACseq; Arabidopsis thaliana; ATAC-seq
library_construction_protocol: Inflorescence tissues were collected for nuclei extraction as follows. About 5 g of inflorescence tissue was collected and immediately transferred into ice-cold grinding buffer (300 mM sucrose, 20 mM Tris pH 8, 5 mM MgCl2, 5 mM KCl, 0.2% Triton X-100, 5 mM β-mercaptoethanol, 35% glycerol). The samples were ground with Omni International General Laboratory Homogenizer at 4 °C and then filtered through a two-layer Miracloth and a 40-µm nylon mesh Cell Strainer (Fisher). Samples were spin filtered for 10 min at 3,000 g, the supernatant was discarded, and the pellet was resuspended with 25 ml of grinding buffer using a Dounce homogenizer. The wash step was repeated once and the nuclei were resuspended in 0.5 ml of freezing buffer (50 mM Tris pH 8, 5 mM MgCl2, 20% glycerol, 5 mM β-mercaptoethanol).  Hi-C libraries for Illumina sequencing were generated53. The libraries were sequenced on HiSeq 2000 DNA sequencers obtaining paired-end 50- or 51-nucleotide reads following manufacturer instructions (Illumina). Nuclei were subjected to transposition reaction with Tn5 (Illumina, cat. no. FC-121-1030). For transposition reaction, 25 µl of 2x DMF (66mM Tris-acetate (pH=7.8), 132 mM K-Acetate, 20mM Mg-Acetate, 32 % DMF) was mixed with 2.5 µl Tn5 and 22.5 µl nuclei suspension at 37 °C for 30 min. Transposed DNA fragments were purified with ChIP DNA Clean & Concentrator Kit (Zymo, cat. no. D5205). Libraries were preparated with Phusion High-Fidelity DNA Polymerase (NEB, cat. no. M0530S) (12.5µl 2x Phusion, 1.25µl 10 mM Ad1 primer, 1.25µl 10 mM Ad2 primer, 4 µl ddH2O and 6µl purified transposed DNA fragments).
identifiers.namespace: GEO
identifiers.id: GSM4705423
attributes.tag: GEO Accession
attributes.value: GSM4705423