title: Replicate 3 Raji iBZLF1 full-length low
organism: Homo sapiens
description: Protocols: For RNA sequencing parental Raji cells and their derivates carrying the conditional expression plasmids encoding full-length iBZLF1 or AD-truncated iBZLF1 were employed. At an initial cell concentration of 5×105 cells/ml, a total of 2×107 non-induced cells or 4×107 cells induced with 100 ng/ml doxycycline for 6 h were analyzed. Doxycycline leads to the co-expression of BZLF1 and the truncated human NGF-receptor. To limit the analysis to BZLF1 expressing cells, only, we sorted NGF-R-positive cells with magnetic beads (MACS, Miltenyi Biotec) and the primary anti-NGF-R antibody (HB8737-1, IgG1) and a secondary anti-mouse IgG (1:10, Miltenyi Biotech) antibody. 7.5×105 cells were lysed in 1 ml Trizol (Thermo Fisher Scientific), snap frozen in liquid nitrogen and stored at −80 °C. Prior to RNA isolation identical molar amounts of ERCC spike-in control RNAs (Ambion) were added to 3×105 non-induced or doxycycline induced Raji iBZLF1 (0h/6h) cells. The subsequent processes and steps were identical for all cells. RNA was extracted with the Direct-zol RNA MiniPrep Kit (Zymo Reseach). The RNA concentration and quality were controlled with Nanodrop (Thermo Fisher Scientific), Quibit (Thermo Fisher Scientific), and Bioanalyzer (Agilent), and the RNAs were treated with dsDNase (Thermo Fisher Scientific). The Encore Complete RNA-Seq Library System kit (NuGEN), which uses not-so-random hexamer primers for rRNA depletion, was used for library preparation. The samples were sequenced on an Illumina HiSeq 1500 instrument (100 nt, single-end reads).
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2019-04-05T04:02:49Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2019-03-26T22:19:24Z
attributes.tag: External Id
attributes.value: SAMEA5537248
attributes.tag: INSDC center name
attributes.value: Research Unit Gene Vectors Helmholtz Zentrum Munchen German Research Center for Environmental Health and German Center for Infection Research (DZIF), Partner site Munich, Germany
attributes.tag: INSDC first public
attributes.value: 2019-04-05T04:02:49Z
attributes.tag: INSDC last update
attributes.value: 2019-03-26T22:19:24Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-7820:Replicate 3 Raji iBZLF1 full-length low
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell line
attributes.value: Raji
attributes.tag: cell type
attributes.value: B cell
attributes.tag: common name
attributes.value: human
attributes.tag: disease
attributes.value: Burkitts lymphoma
attributes.tag: genotype
attributes.value: doxycycline-inducible full-length BZLF1
attributes.tag: organism part
attributes.value: blood
attributes.tag: sample name
attributes.value: E-MTAB-7820:Replicate 3 Raji iBZLF1 full-length low
attributes.tag: scientific_name
attributes.value: Homo sapiens