scRNA Seq of sus scrofa: FMDV A12 W105A infected SK6 cells
SK6 cells were infected with either A12 WT, A12W105A, or A12-LLV at a MOI of 5. After 1 hour of adsorption of virus, cells were treated with MES buffer to inactivate any virus that had not entered the cells. The cells were then incubated for an additional 4 hours at 37C 5% CO2. Following incubation, the cells were dissociated from the flasks using TrypLE (Gibco, Life Technologies Corporation, Grand Island, NY) centrifuged at 1200 x g, and washed three times with buffer (Phosphate-buffered saline (PBS)) containing 2% FBS to create a single-cell suspension. Finally, the cells were counted using a hemacytometer with trypan blue staining to assess viability. Libraries were prepared from single cell suspensions (aimed to recovery 10,000 cells) using 10x Genomics Chromium Single Cell 3 Reagents Kit v3.1 (Cat: 1000691) (10x Genomics, Pleasanton, CA) following manufacturers protocol. Samples were sequenced using NextSeq550 (Illumina, San Diego, CA) following the run parameters: read 1 for 28 cycles, i7 index for 10 cycles, i5 index for 10 cycles, and read 2 for 90 cycles.