characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2014-12-17T17:02:21Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T20:11:50Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2646272
characteristics.characteristic: External Id
characteristics.text: EBI
characteristics.characteristic: INSDC center name
characteristics.text: 2014-12-17T17:02:21Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T20:11:50Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS506251
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-2700:iCLIP_GFP-A3G_CEM-SS_HIV_Producer_Cell_Hs_NNNAATANN_20110921_LUe39_6
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: CEM-SS
characteristics.characteristic: cell line
characteristics.text: T cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0000084
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: CEM-SS T cell lines were grown in RPMI media supplemented with foetal bovine serum and Pen/Strept. HIV-1 viruses were produced from CEM-SS T cell lines by infecting cells with delta Vif IIIB strain HIV-1. HIV-1 viruses produced from CEM-SS T cell lines over expressing proteins of interest were concentrated and purified by ultracentrfugation through a sucrose gradient. Producer cells were washed throughly. Viruses or cells were then ressuspended in PBS and subjected to UV-C irradiation. Cells or viruses were ressuspended in 1mL of lysis buffer (50mM Tris-HCL, pH 7.4; 100mM NaCl; 1% NP-40; 0.1% SDS; 0.5% Sodium deoxycholate and protease inhibitor) and sonicated. 0.04ng of RNase A were added to the samples. RNAs cross-linked to proteins were extracted by immunoprecipitation of the proteins of interest iCLIP protocol was followed to isolate RNA bound to proteins of interest and amplify cDNA library. Proteins of interest were immunoprecipitated using anti-T7 or anti-GFP antibodies.. In order to differentiate between identical reads generated by PCR or distinct reads that were immunoprecipitated but that have the same sequence and length , we inserted 2 random nuclotides 5 prime of the barcode and 3 random nucleotides 3 prime of the barcode on each of the primers used to generate cDNA. Reads that have the same random nucleotides were judged to be artifacts from the PCR. This approach has been described in detail in Konig, et al, (2010): iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution, Nat Struct Mol Biol 17:909-915.. The final products were obtained by PCR with Illumina pair-end sequencing primers: 5-CAAGCAGAAGACGGCATACGAGATCGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATCT-3; 5-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT-3
characteristics.characteristic: description
characteristics.text: GFP-A3G overexpression
characteristics.characteristic: genotype
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: fresh specimen
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/OBI_0000971
characteristics.characteristic: specimen with known storage state
characteristics.text: iCLIP_GFP-A3G_CEM-SS_HIV_Producer_Cell_Hs_NNNAATANN_20110921_LUe39_6
characteristics.characteristic: title