BioProject

We immunosequenced peptide-stimulated PBMCs from 8 CMV-positive donors to identify virus-specific T cell receptors. More...

We immunosequenced peptide-stimulated PBMCs from 8 CMV-positive donors to identify virus-specific T cell receptors. We also sequenced ex vivo T cell repertoires of multiple CMV-postive and CMV-negative donors to compare the availability of CMV-specific TCRs in virus carriers and uninfected subjects. Overall design: Blood samples from healthy anonymous donors were obtained from the "Institut für Transfusionsmedizin Ulm ", Germany. PBMCs were isolated by standard Ficoll gradient centrifugation and mRNA was isolated either directly or after 10-day stimulation with an immunogenic CMV peptide. For validation of our short-term stimulated TCRs, PBMCs from 3 different donors were stimulated with autologous transformed B cells (mini-LCLs) for 23 or 30 days. These mini-LCLs expressed one of the CMV antigens pp65 or IE1 together with EBV antigens, or EBV antigens alone (control mini-LCLs). TCR-enriched sequencing libraries were obtained using TCR beta chain-specific primers with Illumina adapter sequence overhangs. After Sequencing on an Illumina HiSeq1500 device, raw data was demultiplexed, quality filtered, and TCR repertoires were generated by alignment and grouping of individual reads. Less...