GSM5506059: Mouse, adult, male, knockout 76 [snATAC-Seq]; Mus musculus; ATAC-seq
The right kidney was excised, decapsulated, cut into 5 mm cubes and snap frozen. Nuclei were isolated with Nuclei EZ Lysis buffer (NUC-101; Sigma-Aldrich) supplemented with protease inhibitor (cOmplete™ ULTRA tab; Sigma-Aldrich) . The frozen kidney cubes were placed in a sterile petri dish with 1 ml of Nuclei EZ lysis buffer (plus protease and RNase inhibitors) and quickly minced.  The tissue was then homogenized with RNase-free, disposable pellet pestles and tubes (Fisher #12-141-368), and a hand held, Bel-Art™ Micro-Tube homogenizer (Wayne, NJ).  The homogenate was filtered through a 40 micron strainer (all strainers were purchase from pluriSelect Life Science).  The homogenate was moved to a sterile, RNase-free 15 ml tube with an additional 2 ml of Nuclei EZ lysis buffer (plus protease and RNase inhibitors).  After a 5 min incubation on ice, the homogenate was filtered through a 40 micron strainer.  The subsequent homogenate was centrifuged at 500 g for 5 min at 4°C.  The pellet was resuspending in Nuclei EZ lysis buffer (plus 1 ml/ml of SUPERaseIN and RNAsin), incubated on ice for 5 min and centrifuged again.  The pellet was resuspended in 0.5 ml of  10X Genomics 1X nuclei suspension buffer, filtered through a 5 micron strainer, and nuclei immediately counted with a hemocytometer. 4000 nuclei per sample were loaded in the Chromium Next GEM Chip H Single Cell Kit and the DNA libraries generated with the Chromium Next GEM Single Cell ATAC Library & Gel Bead Kit v1.1