title: Positive regulators of IFN lambda signaling
organism: Homo sapiens
description: Protocols: HAP1 cells were derived from the near-haploid chronic myeloid leukaemia cell line KBM7 as described earlier (Carette et al., 2011). HAP1 cells were cultured in IMDM supplemented with 10% FBS, penicillin-streptomycin and L-glutamine. Cells were mutagenized using a RFP gene-trap virus as described in the article with PMCID number: PMC3111863 Positive regulators of IFN lambda signaling: HAP1 cells were transduced with the vector 5xISRE-eGFP to create an ISG reporter cell line. The reporter cell line was sensitized to IFNλ by overexpressing IFNLR1 (IL28RA). These cells were then sub-cloned to obtain a clonal population prior to mutagenesis. An RFP-gene trap virus was used to create a mutagenized HAP1 reporter library. 100 million mutagenized cells were stimulated with 10 IU/ml IFNλ2. After 24 h, stimulated cells were sorted for GFP-negative cells (approximately 5% of the population) and grown for 3 days. The cells were stimulated again with 10 IU/ml IFNλ2 for 24 h and sorted on GFP-negative cells to enrich for cells that no longer responded to IFN treatment. Thirty million cells of the selected population were used for genomic DNA isolation. Genomic DNA was harvested from 30 million cells using the QIAamp DNA mini kit (Qiagen 51304) Sequencing libraries of genomic regions flanking gene-trap integration sites in control and sorted cell populations were prepared using a linear amplification (LAM) PCR-mediated method described in the article with PMCID number: PMC3111863
attributes.tag: ENA first public
attributes.value: 2017-07-25
attributes.tag: ENA last update
attributes.value: 2017-07-11
attributes.tag: External Id
attributes.value: SAMEA104158623
attributes.tag: INSDC center alias
attributes.value: STANFORD UNIVERSITY
attributes.tag: INSDC center name
attributes.value: Stanford University
attributes.tag: INSDC first public
attributes.value: 2017-07-25T17:01:20Z
attributes.tag: INSDC last update
attributes.value: 2017-07-11T12:20:18Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-5866:Positive regulators of IFN lambda signaling
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell line
attributes.value: HAP-1
attributes.tag: cell type
attributes.value: haploid cell
attributes.tag: common name
attributes.value: human
attributes.tag: disease
attributes.value: chronic myelogenous leukemia
attributes.tag: genotype
attributes.value: ISRE-GFP reporter and IL28RA overexpression
attributes.tag: phenotype
attributes.value: GFP-negative
attributes.tag: sample name
attributes.value: E-MTAB-5866:Positive regulators of IFN lambda signaling