Single nuclei RNA sequencing of patient sample with 5' 10X primers: Sample UM_sc_5p
Tissue specimens were kept in ice cold RPMI in a petri dish and cut into 1 mm3 cubes using two scalpels. The cubes and RPMI were transferred to a 50 ml Falcon tube (Corning) using a 10 ml serological pipette and collected by 5 min centrifugation at 300 x g at 4°C. The tissue was then digested using human tumor dissociation enzymes (Miltenyi Human Tumor Dissociation Kit; #30-095-929) according to manufacturer instructions based on the tissue weight. Samples were loaded on a Chromium controller with Chromium Single Cell V(D)J Reagents (10X Genomics, Pleasanton, CA; #1000006) for 5' RNA capture. Gene expression libraries were then generated using Chromium Single Cell 5' Library construction kit (#1000020) according to manufacturer instructions.