Single Cell of Bos taurus: PBMCs
PBMCs libraries were prepared from single cell suspensions of 10,000 cells using 10x Genomics Chromium Single Cell 3 Reagents Kit v3.1 (Cat#: 1000691) (10x Genomics, Pleasanton, CA) following manufacturers protocol. The single cell suspension, gel beads and oils were added to the 10x Genomics single-cell G chip. After droplet generation, reverse transcription was performed on samples using a Mini Amp thermocycler (Applied Biosystems, Rockville, MD). After reverse transcription, cDNA was recovered using a recovery agent provided by 10x Genomics, followed by Silane DynaBead clean-up as described in the manufacturers manual. Before clean-up using SPRIselect beads, we amplified the cDNA for 12 cycles. Library quality was determined on Qubit Flex (Invitrogen, Life Technologies, Carlsbad, CA). The cDNA libraries were then prepared according to the Chromium Single Cell 3 Reagent Kit v2 user guide. Quality of libraries were determined on Qubit Flex and Agilent TapeStation 4200 System (Agilent, Santa Clara, CA). Samples were sequenced by NextSeq 550 (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.