Experiment Detail
| Title | Illumina HiSeq 1500 sequencing; RNA-seq in Raji cells with inducible BZLF1 prior to and after induction of EBV's lytic cycle by doxycycline |
| Design Description | RNA-seq in Raji cells with inducible BZLF1 prior to and after induction of EBV's lytic cycle by doxycycline |
| Organism | Homo sapiens |
Library Description
| Name | Replicate 2 Raji iBZLF1 full-length low_s |
| Strategy | RNA-Seq |
| Source | TRANSCRIPTOMIC |
| Selection | Inverse rRNA |
| Layout | SINGLE |
| Construction Protocol | 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). |
Platform
| Platform | ILLUMINA |
| Instrument Model | Illumina HiSeq 1500 |
Processing
| Base Calls | |
|---|---|
| Sequence Space | |
| Base Caller | |
Spot Information
| Number of Reads per Spots | 0 |
| Spot Length | 0 |
Read Spec
Navigation
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Submission | ERA1800239 |
 FTP
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Study | ERP114480 | ||
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Sample | ERS3339307 | ||
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Run | ERR3252902 |
FASTQ
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SRA
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