title: GSM8545120: Day30Pool_rep1_GEX; Mus musculus; RNA-Seq
library_construction_protocol: live CD8β+CD44hiCD45.1+Vα2+ P14 cells from each sample (pooled organs from 20 Arm-infected mice or 20 C13 infected mice) were sorted into separate FBS-coated FACS tubes (Corning) containing 50%RPMI/FBS samples were stained with individual TotalSeq-A Hashtag Oligo (HTO) antibodies and a custom pool of TotalSeq-A Antibody Derived Tag (ADT) antibodies (Biolegend; see Supplementary Table X). Samples were washed with Cell Staining Buffer (850G, 7 mins, 4°C) before resuspending cells in 2%BSA/DPBS and pooling cells. Cells were resuspended in custom TEA-seq Wash Buffer (20mM Tris-HCl, 150mM NaCl, 2mM MgCl2 and 1% BSA in H2O) containing RNase Inhibitor (1U/mL, Roche) and 0.01% Digitonin (Thermofisher) and incubated for 5 minutes on ice to permeabilize cell and nuclear membranes. Cells were topped with TEA-seq Wash Buffer and spun for 7 mins at 850G at 4°C before washing with TEA-seq Tagmentation Buffer (20mM Tris-HCl, 150mM NaCl, 2mM MgCl2 in H2O). 30,000 cells per reaction were removed and subjected to transposition and GEM generation as described in the 10x Chromium Single Cell Multiome ATAC + Gene Expression Kit across a total of two wells. Library preparation was performed as described47, with the following customizations. Indexing of ADT and HTO libraries was performed using Biolegend-recommended Illumina D70x (i7 long; HTO) or RNA PCR Index (RPIx short; ADT) adapter sequences (IDT), with three additional rounds of amplification performed on purified HTO/ADT libraries using P5/P7 primers specific for generic Illumina adapter sequences (IDT) prior to sequencing