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Preprint

Single-cell RNA-seq analysis of public 10x data

JG Jean-Baptiste Gorin
VW Vincent Wu
SN Son Nguyen
MB Marcus Buggert

Last updated date: Dec 7, 2020 Views: 848 Forks: 0

original An abbreviated version of this protocol was published in Science Immunology
TOX is expressed by exhausted and polyfunctional human effector memory CD8+ T cells
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1. Download data


scRNA-seq data were obtained from the public 10× genomics application note: A New Way of Exploring Immunity (www.10xgenomics.com/resources/application-notes/a-new-way-of-exploring-immunity-linking-highly-multiplexedantigen-recognition-to-immune-repertoire-and-phenotype/).

  • donor 1
    $ curl -O https://cf.10xgenomics.com/samples/cell-vdj/3.0.2/vdj_v1_hs_aggregated_donor1/vdj_v1_hs_aggregated_donor1_filtered_feature_bc_matrix.tar.gz

  • donor 4
    $ curl -O https://cf.10xgenomics.com/samples/cell-vdj/3.0.2/vdj_v1_hs_aggregated_donor4/vdj_v1_hs_aggregated_donor4_filtered_feature_bc_matrix.tar.gz


2. Normalize and integrate


CD8+ T cell gene expression data from both healthy donors was then normalized with SCTransform and integrated with Seurat v3 using the script integrate_d1_d4.R available on GitHub: https://github.com/Jb-Gorin/Sekine-et-al.-2020/blob/master/integrate_d1_d4.R


3. DEG

Tox+ (1909) and Tcf+ (9430) cells were identified, and differentially expressed genes (DEGs) were determined using DESeq2 using the script DEG_Tox_Tcf7.R available on GitHub: https://github.com/Jb-Gorin/Sekine-et-al.-2020/blob/master/DEG_Tox_Tcf7.R 

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How to cite：
Readers should cite both the Bio-protocol preprint and the original research article where this protocol was used:
  1. Gorin, J, Wu, V, Nguyen, S and Buggert, M(2020). Single-cell RNA-seq analysis of public 10x data. Bio-protocol Preprint. bio-protocol.org/prep683.
  2. Sekine, T., Perez-Potti, A., Nguyen, S., Gorin, J., Wu, V. H., Gostick, E., Llewellyn-Lacey, S., Hammer, Q., Falck-Jones, S., Vangeti, S., Yu, M., Smed-Sörensen, A., Gaballa, A., Uhlin, M., Sandberg, J. K., Brander, C., Nowak, P., Goepfert, P. A., Price, D. A., Betts, M. R. and Buggert, M.(2020). TOX is expressed by exhausted and polyfunctional human effector memory CD8+ T cells . Science Immunology 5(49). DOI: 10.1126/sciimmunol.aba7918

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