GSM6807112: Atoh7-zsGreen-wt-E14.5 scRNA; Mus musculus; RNA-Seq
Nuclei isolation was performed following the protocol from 10X Genomics (CG000169). Briefly, 100,000-1,000,000 cells were collected by FACS and centrifuged at 300 g for 5 min at 4°C. Cell pellets were resuspended in 100 μl cold Lysis Buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 0.1% Nonidet P40, 0.01% Digitonin, 1%BSA), mixed by gently by pipetting then incubated for 2 min on ice. One milliliter of cold Wash Buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 1%BSA) was then added immediately and mixed by gentle pipetting. The cells were then spun down at 500 g for 5 min at 4°C, and resuspended in cold 1X Nuclei Buffer (10x Genomics, PN-2000153). The nuclei concentration and quality were determined with a hemocytometer. scRNA-seq:  scRNA-seq was performed on 10X Chromium single cell platform using the Chromium Single Cell 3' Library & Gel Bead Kit v3.1, in keeping with the manufacturer's instructions (10X Genomics, CG000204). The sorted cells were loaded onto the 10X Genomics Chromium Controller targeting >5,000 cells per sample to generate scRNA-seq libraries. The quantity and quality of the libraries were evaluated by the Qubit Fluorometer and Agilent Fragment Analyzer. High-quality libraries were subsequently sequenced on a NovaSeq6000 sequencer (PE 26X100). snATAC-seq:  scATAC-seq library preparation was performed following the protocol from 10X Genomics (CG000168 Rev B) using the Chromium Single Cell ATAC Reagent Kits (10X Genomics). The libraries were cleaned using SPRIselect reagent (Beckman Coulter). The quantity and quality of the libraries were evaluated by the Qubit Fluorometer and Agilent Fragment Analyzer. The libraries were subsequently sequenced on a NovaSeq6000 (PE50). CUT&Tag:  The Cleavage Under Targets & Tagmentation (CUT&Tag) experiments were performed using reagents and following a protocol from EpiCypher. scRNA-Seq (10 Genomics), snATAC-Seq (10 Genomics), CUT & Tag (Epicypher)