organism: Mus musculus
description: Protocols: 46c mouse ES cells were grown feeder-free on 0.2% gelatinized cell culture plates in 2i-media. The 2i-media contained DMEM/F12 media for SILAC (Pierce, 88215) supplemented 100 mg/ml of Lysine (Sigma, L8662), 100 mg/ml of Arginine (Sigma, A6969) and 100 mg/ml of Proline (Sigma, P5607), 100 microM MEM non-essential amino acids (Life technologies, 11140-050), 1xGlutamax (Life technologies, 35050-061), 1x penicillin and streptomycin (Life technologies, 15140-122), 100 microM of 2-mercaptoethanol (Sigma, M7522), 0.5mg/ml of BSA (Sigma, A3059), 200ng/ml of LIF (EMBL, protein expression core facility), 1microM of PD0325901(Reagents Direct, 39-C68), 3 microM of CHIR99021 (Reagents Direct, 27-H76). The cells were cultured using the 2i-media at least for 10 days, and passaged 4 times. The knock down (KD) was carried out by lentiviral transduction of : non-targeting shRNA, shTrim24 (TRCN0000088518), shTrp53 (TRCN0000310844), and the mixture of the last two shRNAs. For each knock down, 3 independent transductions was perfomed. For the details of virus production and infection please refer to PMID: 23260666. 48 hours after the viral treatment of the mouse ES cells, the cells were lysed, and RNA was extracted to be analyzed by qPCR and mRNA-seq RNA was extracted by Qiagen kit (RNeasy mini kit) mRNA-Seq libraries (from polyA+ cytoplasmic RNA) were created according to the standard Illumina TruSeq protocol.
attributes.tag: ENA first public
attributes.value: 2016-11-08
attributes.tag: ENA last update
attributes.value: 2016-07-04
attributes.tag: External Id
attributes.value: SAMEA4062609
attributes.tag: INSDC center alias
attributes.value: EMBL
attributes.tag: INSDC center name
attributes.value: European Molecular Biology Laboratory
attributes.tag: INSDC first public
attributes.value: 2016-11-08T17:11:37Z
attributes.tag: INSDC last update
attributes.value: 2016-07-04T12:10:51Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-4893:T24KD_Trim_p53_R1
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell line
attributes.value: 46c ES
attributes.tag: common name
attributes.value: house mouse
attributes.tag: genetic modification
attributes.value: transfection
attributes.tag: genotype
attributes.value: Trim24/Trp53 Knock Down
attributes.tag: sample name
attributes.value: E-MTAB-4893:T24KD_Trim_p53_R1