characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-06-01T00:15:25Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-06-01T00:15:25Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA111468771
characteristics.characteristic: External Id
characteristics.text: Faculty of Biology, Medicine and Health, University of Manchester
characteristics.characteristic: INSDC center name
characteristics.text: 2023-06-01T00:15:25Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-06-01T00:15:25Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS13562801
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-12294:ATACseq U2OS
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: U2OS
characteristics.characteristic: cell_line
characteristics.text: human
characteristics.characteristic: common name
characteristics.text: Protocols: Cells were lysed 48 h after transfection in cold lysis buffer (10mM Tris-HCL, pH 7.4, 10mMNaCl, 3mM MgCl2 and 0.1% Igepal). DNA was purified using Qiagen mini elute kit and using Ampure XP beads following the manufacturer’s instructions. Nuclei was concentrated by centrifugation at 500 g for 10 minutes at 4°C. The nuclei pellet was resuspended in nuclease free water, diluted 1:10 and quantified. The equivalent volume for 50,000 nuclei was made up to 22.5 ul with nuclease free water. Nuclei were incubated for 30 minutes at 37°C, 300 rpm, with 25 ul of 2x TD buffer and 2.5 ul tagmentation ezyme (Illumina transposase FC-121-1030). Library fragments were amplified using standard PCR and Nextera primers (Illumina) by adding 25 ul of 2x NEBnext master mix, 2.5 ul forward primer, 2.5 ul reverse primer and 20 ul transposased DNA. PCR was run for 5 cycles, 72°C, 5 minutes; 98°C 30 seconds; cycle of 98°C 10 seconds, 63°C 30 seconds, 72°C 1 minute, paused at 4°C. To determine the total number of cycles needed for the amplified library, 5 ul of the PCR mix were taken and added to a new mix containing 5 ul 2X master mix, 1 ul of each primer, 0.6 ul of 10X SYBR green and 2.4 ul H2O. In a qPCR machine, 20 cycles were run, the total number of cycles for the ATAC library was chosen according to qPCR cycle that gave 1/3 of saturation.
characteristics.characteristic: description
characteristics.text: juvenile stage
characteristics.characteristic: developmental stage
characteristics.text: osteosarcoma
characteristics.characteristic: disease
characteristics.text: wild type genotype
characteristics.characteristic: genotype
characteristics.text: not applicable
characteristics.characteristic: isolate
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: tibia
characteristics.characteristic: organism part
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: ATACseq U2OS
characteristics.characteristic: title