characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-02-17T16:22:12Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-02-17T16:22:12Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA110375049
characteristics.characteristic: External Id
characteristics.text: EMBL
characteristics.characteristic: INSDC center name
characteristics.text: 2023-02-17T16:22:12Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-02-17T16:22:12Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS12473245
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-11993:antiBEAF_WT_Depletion_withSpikeIn_2
characteristics.characteristic: Submitter Id
characteristics.text: 2h10-2h40
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: fruit fly
characteristics.characteristic: common name
characteristics.text: Protocols: Wildtype Drosophila melanogaster (yellow white - yw) flies or crosses for insulator depletions were grown in population cages at 25 degr. Independent collections of staged embryos were performed for each biological replicate. The collected embryos were dechorionated using 50% bleach and formaldehyde fixed in 700 ul cross-linking solution (50 mM Hepes, 1 mM EDTA, 0.5 mM EGTA, 100 mM NaCl, 1.8 % formaldehyde, pH 8.0) and 30ml n-heptane on a shaker table at RT for 15 minutes. Specifically for CP190 C&Ts, the same collection procedure was used, but using a final 3.0 % concentration of formaldehyde, for 30 minutes. The reaction was terminated with 125 mM glycine, 0.1% Triton X-100 in PBS. After washing out the fix, the embryos were blotted dry and frozen in liquid nitrogen. A small number of embryos from each collections was set aside to stage each collection. Nuclei Preparation Step: Nuclei were extracted from the frozen embryos by Dounce homogenization and differential centrifugation (Cell lysis buffer: 5mM Hepes, pH 8, 85mM KCl, 0.5% NP40 + protease inhibitors). The nuclei were counted at a flow cytometer, divided in 50K aliquots and snap-frozen in liquid nitrogen. Antibody Binding and Tagmentation Step: 50K nuclei from D. Melanogaster and D. Virilis were thawed and mixed together on ice. ConA beads were then added to the nuclei mixture. The nuclei were permeabilized, and then the nuclei-beads complex was incubated with the primary antibody and slowly rotated (5 rpm) overnight at 4C. A secondary antibody solution (1:100, 100 _l/sample) was added to the beads and incubated for 1h at room temperature. Samples were then washed three times and incubated with a pA-Tn5-oligo adapter complex. Tagmentation allowed to happen at 37C for 1h. Samples were de-crosslinked, digested with protease, phenol/chloroform - extracted, column purified and RNase treated. C&T NGS libraries were prepared using P5 and P7 primers and the NEBNext� High-Fidelity 2x PCR Mater Mix (M0541L).
characteristics.characteristic: description
characteristics.text: NC14
characteristics.characteristic: developmental stage
characteristics.text: transgenic
characteristics.characteristic: genetic modification
characteristics.text: wild type genotype
characteristics.characteristic: genotype
characteristics.text: egg laying
characteristics.characteristic: initial time point
characteristics.text: Drosophila melanogaster
characteristics.characteristic: organism
characteristics.text: Drosophila melanogaster
characteristics.characteristic: organism
characteristics.text: whole organism
characteristics.characteristic: organism part
characteristics.text: Drosophila melanogaster
characteristics.characteristic: scientific name
characteristics.text: mixed
characteristics.characteristic: sex
characteristics.text: yw
characteristics.characteristic: strain
characteristics.text: antiBEAF_WT_Depletion_withSpikeIn_2
characteristics.characteristic: title