Paired Vgamma9Vdelta2 T-cell receptor sequences from pigtail macaques (Macaca nemestrina) following in vivo pharmacological expansion
Description
Dataset related to the following manuscript: -Barber-Axthelm IM, Wragg KM, Esterbauer R, Amarasena TH, Barber-Axthelm VRB, Wheatley AK, Gibbon AM, Kent SJ, and Juno JA. Phenotypic and functional characterization of pharmacologically expanded Vgamma9Vdelta2 T-cells in pigtail macaques (2023) The included datasets are related to in vivo pharmacologically expanded Vgamma9Vdelta2 T-cells in pigtail macaques (Macaca nemistrina). Gamma and delta T-cell receptor (TCR) sequences were taken from peripheral blood Vgamma9Vdelta2 T-cells collected per-expansion (day -14) or at peak expansion (day 4), from 4 animals (NM11, NM89, NM251, NM295). Sequencing data from Day 15 is also included from 1 animal (NM89). Sequences were amplified by single-cell, multiplex RT-PCR, prior to capillary electrophoresis sequencing. Details on RT-PCR amplification conditions and primers can be found in the associated manuscript. The sequences were subsequently aligned using the MiXCR software package (v3.0.13; species: Macaca mulatta). Dataset description: -Combined_TCR_sequence_alignments_MiXCR.csv: Aligned, paired, single-cell TCRG and TCRD sequences, with associated FACS sort data (from BD FACSAria III cell sorter), for 4 animals (NM11, NM89, NM251, NM295) and 2 timepoints (pre-expansion and peak expansion). Sequencing data from Day 15 is also included from 1 animal (NM89). Rows without TCRG/TCRD alignment information indicate an RT-PCR sequencing product that could not be aligned. Initial RT-PCR TCRG/TCRD nucleotide sequences are included. -TCRG_Functional.clones.csv: Assembled list of functional TCRG clones recovered across 4 animals (NM11, NM89, NM251, NM295) and 2 timepoints (pre-expansion and peak expansion), generated through the MiXCR assemble function. TCRG sequences from Day 15 is also included from 1 animal (NM89). TCRG clone list is used to filter out non-functional clones from the Combined_TCR_sequence_alignments_MiXCR.csv in R. -TCRD_Functional.clones.csv: Assembled list of functional TCRD clones recovered across 4 animals (NM11, NM89, NM251, NM295) and 2 timepoints (pre-expansion and peak expansion), generated through the MiXCR assemble function. TCRG sequences from Day 15 is also included from 1 animal (NM89). TCRD clone list is used to filter out non-functional clones from the Combined_TCR_sequence_alignments_MiXCR.csv in R. The code used for downstream TCR sequence analysis has been deposited at GitHub (https://github.com/BarberAxthelm/Vd2-Vg9_TCR_Analysis) and is publicly available as of the date of publication.
Files
Steps to reproduce
Upstream sequence alignments: -TCRG and TCRD sequence alignments were completed with the raw nucleotide sequences using the MiXCR software package (v3.0.13; species: Macaca mulatta). -Single cell alignments were generated by: 1) align (verbose), followed by 2) exportAlignments. -Aligned TCRG and TCRD sequences were subsequently matched with the corresponding sample information (animal ID, day, PCR plate #, well ID) and FACS data. -Assembled TCRG and TCRD clone lists were generated from combined, aligned clone lists by: 1) assemble, followed by 2) exportClones (excluding out-of-frame clones and sequences with premature stop codons). Downstream analysis: -The code utilized for dowstream TCR analysis has been deposited at GitHub (https://github.com/BarberAxthelm/Vd2-Vg9_TCR_Analysis) and is publicly available as of the date of publication. -An initial dataset of paired, functional, TCRG and TCRD clones is generated from the Combined_TCR_sequence_alignments_MiXCR.csv datafile, using the TCR_Data_Preparation.R code (https://github.com/BarberAxthelm/Vd2-Vg9_TCR_Analysis), with the TCRG_Function.clones.csv and TCRD_Functional.clones.csv data files. -The filtered dataset is then used for analysis and data visualization, using additional code (TCR_CDR3_Spectratyping.R, TCR_Diverstiy.R, TCR_Randomization_Statistics.R, TCR_Alluvial_Plot.R, TCR_Circos_Plots.R, TCR_MSA_Sequence_Logos.R, TCR_Upset_Plot.R, TCRG_CDR3_Shared_Clonotypes.R; https://github.com/BarberAxthelm/Vd2-Vg9_TCR_Analysis) Refer to the README.md for additional information on using the analysis code.