characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2015-12-31T17:01:13Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T19:30:14Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2707069
characteristics.characteristic: External Id
characteristics.text: Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University
characteristics.characteristic: INSDC center name
characteristics.text: 2015-12-31T17:01:13Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T19:30:14Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS528881
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-2840:IP_F_HCT116_3
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: HCT116
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0002824
characteristics.characteristic: cell line
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: RNA immunoprecipitation was performed in HCT116 cells transiently transfected with pCMVtag2b or pCMVtag2bhWig-1 and in Saos-2 TetON cells without insert or Saos-2 TetON cells stably transfected with either Flag-tagged wt Wig-1 or a Flag-tagged Wig-1 zinc-finger 1 point mutant that cannot bind to RNA. Transfected cells were lysed in RIP buffer B (150 mM KCl, 25 mM Tris-HCl, pH 7.5, 10 uM ZnCl2, 1% NP40, 1% Protease Inhibitor Cocktail, 0.5 mM DTT, 100 U/mL RNaseOUT), cleared by centrifugation at 20,000g 30 min at 4C, and filtered through a 45um filter. Lysates containing 30mg of protein were subjected to IP for 2 h rotating at 4C on 600 uL of anti-FLAG M2 beads, pre-equilibrated three times with RIP buffer A (150 mM KCl, 25 mM Tris-HCl, pH 7.5, 10 uM ZnCl2) and once with RIP buffer B rotating at 4 C for 10 min. Input was removed, and beads were washed twice with buffer B rotating at 4 C for 10 min, collected between washes by centrifugation at 50g for 2 min at 4C. RNA was extracted by two rounds of phenol/chloroform/isoamyl alcohol and one round of chloroform extraction, each round consisting of three times 30s vortexing and 30s incubation at 60C, followed by centrifugation at 20,000g for 5 min. RNA was precipitated by using 1 vol of isopropyl alcohol, 0.1 vol of sodium acetate, and 1uL of glycoblue. RNA libraries for sequencing were prepared using TruSeq RNA Sample Prep (Illumina, CA, USA).
characteristics.characteristic: description
characteristics.text: 12
characteristics.characteristic: individual
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: IP_F_HCT116_3
characteristics.characteristic: title