characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2022-05-05T00:12:40Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2022-05-05T00:12:40Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA12598058
characteristics.characteristic: External Id
characteristics.text: Research Unit Gene Vectors, Helmholtz Zentrum Munchen, German Research Center for Environmental Health and German Center for Infection Research, Munich, Germany
characteristics.characteristic: INSDC center name
characteristics.text: 2022-05-05T00:12:40Z
characteristics.characteristic: INSDC first public
characteristics.text: 2022-05-05T00:12:40Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS10204525
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-11350:18-wt-3
characteristics.characteristic: Submitter Id
characteristics.text: TTGCTCCGCAGA
characteristics.characteristic: barcode
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: B cell
characteristics.characteristic: cell type
characteristics.text: human
characteristics.characteristic: common name
characteristics.text: Protocols: Isolated, primary B cells were isolated from human adenoid or tonsil samples and sorted for naive, resting (CD38-ive/IgD+ive) B cells. 10,000 cells were used as the uninfected control (day 0). The rest was either infected with wild type EBV or mutant EBV with an MOI: 0.1. On day 1, 2, 3, 4 post-infection viable cells were sorted based on the forward and site scatter, counted and 10,000 directly added to RLT+ plus lysis buffer with ß-mercaptoethanol. All samples were stored immediately at -80 °C until the samples of 6 biological replicates were collected. RNA extraction and cDNA library preparation is based on the prime-seq protocol. Lysates were first treated with Proteinase K (Ambion) followed by a beads clean-up (2:1 beads to lysate ratio) using solid phase reversible immobilisation (SPRI) beads (GE Biotech). A subsequent on-bead DNAseI digest was performed to remove genomic DNA, followed by another SPRI bead clean-up and reverse transcription. Reverse transcription (RT) was performed using Maxima H- RT enzyme (Thermo Fisher Scientific) with well specific barcoded oligo dT primers and a template switch oligo (TSO). After the RT, the barcoded first strand cDNA was pooled and cleaned up using magnetic beads. Remaining oligo dT primers were removed using exonucleaseI digest. Subsequently, cDNA was PCR amplified with Kapa Hifi DNA polymerase (Roche), quantified using picogreen dye (Thermo Fisher Scientific) and quality control using capillary gel electrophoresis (Agilent Bioanalyzer 2100). Sequencing libraries were generated with the NEB NEXT Ultra II FS kit by using a custom adapter oligo and a primer, specific for the cDNA 3’- end. Fragments between 300 bp and 500 bp were selected using double size selection with SPRIselect beads (Beckman Coulter). Final libraries were quantified and quality controlled again with the Bioanalyzer 2100.
characteristics.characteristic: description
characteristics.text: CD38 negative, IgD positive
characteristics.characteristic: immunophenotype
characteristics.text: 18
characteristics.characteristic: individual
characteristics.text: Epstein-Barr virus wild type strain
characteristics.characteristic: infect
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: adenoid or tonsil
characteristics.characteristic: organism part
characteristics.text: 5
characteristics.characteristic: replicate
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: 18-wt-3
characteristics.characteristic: title