characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2014-03-03T16:32:09Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T16:41:30Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2177664
characteristics.characteristic: External Id
characteristics.text: University of Rome La Sapienza. Dipartimento di Biotecnologie Cellulari ed matologia, Sezione di Genetica Molecolare
characteristics.characteristic: INSDC center name
characteristics.text: 2014-03-03T16:32:09Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T16:41:30Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS345758
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-1844:cont_rep1
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: HeLaS3
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0002791
characteristics.characteristic: cell line
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: HeLaS3 cells were grown in DMEM supplemented with 10%FBS, pen/strep and L-glutamine. HeLaS3 cells were transfected with 10nMol/l siRNA using Interferin reagent (Polyplus) according to manufacturer instructions. AGO2 reduction was confirmed by Western blot analysis. (1) Pellet 2x 106 cells . (2) Add 300uL of Permeabilization buffer (15mM Tris-HCl pH 7.4, 300mM sucrose, 60 mM KCl, 15 mM naCl, 4mM CaCl2, 0.5mM EGTA, 0.2%NP-40, 0.5mM beta-mercaptoethanol) on ice. (3) Add 50U of Mnase. (4) Incubate for 20 min at 37 degrees C (HeLa S3). (5) Stop reaction by adding 300uL of Stop Buffer (50mM Tris-HCl pH 8, 20mM EDTA, 1%SDS), place the tube on ice and wait 2 min. (6) RNase treatment: add 75uG RNase A 1h at 37 degrees C. (7) Proteinase K treatment: add 30uG RNase A 1h at 55 degrees C. (8) DNA was purified by adding 1 volume of phenol:chloroform:isoamyl alchol, vortex and centrifuge max speed 5-10 min 4 degrees C . Repeat. Add 1 volume of chloroform, vortex and centrifuge max speed 5-10 min 4 degrees C .
characteristics.characteristic: description
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: frozen specimen
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/OBI_0000922
characteristics.characteristic: specimen with known storage state
characteristics.text: cont_rep1
characteristics.characteristic: title