GSE158102
Single cell RNA-seq of Oncorhynchus mykiss B cells isolated from blood
These files represent single cell RNA-Seq data generated on a 10x Chromium genomics platform from Oncorhynchus mykiss B cells isolated from blood. Overall design: Three healthy rainbow trout were killed by benzocaine overdose. To isolate peripheral blood leukocytes (PBLs) blood was drawn with a heparinized needle from the caudal vein and diluted 10 times with of Leibovitz medium. The resulting cell suspension was placed onto 51% Percoll density cushions collecting the interface cells after centrifugation. Leukocytes were incubated with anti-trout MHC-II APC and YO-PRO in FACS staining buffer. MHC-II+ leukocytes from lymphoid gate were isolated by sorting. A total of 2,500 cells (700 cells/µL) from each fish were loaded to chips from the Chromium Single Cell 5' Gel Beads Kit (10x Genomics, CA, USA) and subjected to the Chromium Controller Instrument (10x Genomics) to generate single-cell gel beads in emulsion following manufacturer's instructions. Libraries for sequencing were constructed using Chromium Single Cell 5' Library Construction Kit following manufacturer's instructions. Libraries were sequenced using a NextSeq instrument (Illumina) with 150PE chemistry.