title: GSM8284032: whole spleen, unfractionated, biological replicate 1; Ictalurus punctatus; RNA-Seq
library_construction_protocol: Three channel catfish (~ 1 kg) were euthanized in a solution of buffered MS-222 for 10 min. The individual spleens were removed and placed into Petri dishes containing Leibovitz's L-15 medium adjusted to catfish tonicity (cL-15) with PenStrep glutamine solution and 5% FBS.  The spleens were first processed through 70 µM and then 40 µM cell sieves into a 50 mL Falcon tube and cL-15 was added up to 10 mL. The splenic cell suspensions were washed 3 times in 10 mL cL-15 and centrifuged at 350 x g for 5 min. The final cell pellet was resuspended in 15 mL cL-15. Cells were counted, and cell viability was assessed. Cells were pelleted and resuspended in 1 mL of freezing medium (90% FBS, 10% DMSO) and placed into a Mr. Frosty freezing container at -80°C for 24 h. The cryogenic vials were then moved into a liquid nitrogen storage until shipping to a sequencing vendor. The single-nuclei RNA-seq libraries were prepared and constructed using the Chromium X Instrument (10x Genomics) and the Chromium Next GEM Single Cell 3' GEM Kit v3.1 (10x Genomics) following the manufacturer's protocol.