title: GSM8543511: whole kidney snRNA seq of mouse #4443; Mus musculus; RNA-Seq
library_construction_protocol: Nuclei were extracted from flash frozen whole kidney samples. The specimens were thawed and minced thoroughly for one minute in NP40 lysis buffer (0.1% NP40, 10 mM Tris pH 7.5, 146 mM NaCl, 1 mM CaCl2, 21 mM MgCl2, and 40 U/mL RNAse inhibitor). Next, the suspension was further homogenized in 1 mL lysis buffer using a Dounce homogenizer (Kimble) with 20 strokes each using pestle A and pestle B. Subsequently, 4 mL of wash buffer (10 mM Tris, 146 mM NaCl, 1 mM CaCl2, 21 mM MgCl2, 1% BSA [7.5% Fraction V], and 40 U/mL RNAse inhibitor) was added, filtered through a 30 µm strainer (Milteni), and centrifuged for 10 min at 200×g at 4 °C. The supernatant with debris was carefully removed using a glass Pasteur pipette and house vacuum, and the pellet was then resuspended in 5 mL wash buffer, filtered through a 5 µm strainer (Pluriselect), and centrifuged again.  Nuclei were submitted to the Advanced Genomics Core at the University of Michigan for single-nucleus RNA sequencing using the 10X Genomics Chromium platform, following manufacturer instructions. cDNA libraries were prepared using the Chromium platform following manufacturer instructions (10X Genomics).