Homo sapiens
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 .
ENA-FIRST-PUBLIC
2014-03-03T16:32:09Z
ENA-LAST-UPDATE
2018-03-08T16:41:30Z
External Id
SAMEA2177663
INSDC center name
University of Rome La Sapienza. Dipartimento di Biotecnologie Cellulari ed matologia, Sezione di Genetica Molecolare
INSDC first public
2014-03-03T16:32:09Z
INSDC last update
2018-03-08T16:41:30Z
INSDC status
public
Submitter Id
E-MTAB-1844:AGO2_KD_rep1
broker name
ArrayExpress
cell line
HeLaS3
common name
human
sample name
E-MTAB-1844:AGO2_KD_rep1
scientific_name
Homo sapiens
specimen with known storage state
frozen specimen