GSM6625632: PBS-treated WT; Mus musculus; RNA-Seq
24 hours after infection, mice were euthanized and their lungs were harvested. To create a single cell suspension, the lungs were placed in an Eppendorf tube containing an enzymatic digestion solution of 2 mg/mL collagenase I (Gibco 17100-017), 20 mg/mL dispase (Gibco 17105-041), 1 mg/mL elastase (Worthington LS006365), and 1 uL/mL DNAse (Invitrogen 2224G) in PBS. The lungs were minced in the Eppendorf tube, then incubated with shaking at 37 degrees Celsius for 30 minutes. The digestion was quenched with 4 volumes of PBS supplemented with 10% heat inactivated fetal bovine serum (hiFBS; Sigma F4135), and strained over a 70-micron filter (CELLTREAT 229483). The cell suspension was centrifuged at 1400 xg for 7 minutes at 4°C. Red blood cell lysis was performed with the eBioscience RBC lysis buffer. The resulting cell pellet was resuspended in PBS supplemented with 0.04% bovine serum albumin (BSA; Sigma-Aldrich A7409) before being loaded onto the 10X Genomics Chromium Single Cell Controller. Libraries were constructed according to manufacturer's (10X Genomics) instructions.