title: GSM8941100: snMultiome, PA_001_M, ATAC; Homo sapiens; ATAC-seq
library_construction_protocol: snMultiome: Briefly, and all on ice, RNAse inhibitor (NEB MO314L) was added to the homogenization buffer (0.32 M sucrose, 1 mM EDTA, 10 mM Tris-HCl, pH 7.4, 5mM CaCl2, 3mM Mg(Ac)2, 0.1% IGEPAL CA-630), 50% OptiPrep (Stock is 60% Media from Sigma; cat# D1556), 35% OptiPrep and 30% OptiPrep right before isolation. Each pituitary was homogenized in a dounce glass homogenizer (1ml, VWR cat# 71000-514), and the homogenate filtered through a 40 mm cell strainer. An equal volume of 50% OptiPrep was added, and the gradient centrifuged (SW41 rotor at 9200rpm; 4C; 25min). Nuclei were collected from the interphase, washed, resuspended in 1X nuclei buffer for sn multiome (10X Genomics). snMultiome: sn Multiome (10x Genomics, Pleasanton, CA) was performed  following the Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Kits User Guidelines. Nuclei were filtered and counted on a Nexcelom instrument. The amount of nuclei loaded on the chip was determined based on the 10X Genomics protocol loading table. The 10X chip (Chromium Single Cell Chip kit J  PN-200264) was loaded to target 2,000-10,000 nuclei final. After transposition, linear amplification was performed in the emulsion and cDNA was amplified before the RNA and ATAC library constructions were performed following the Chromium multiome protocol. The library was tagged with an index for multiplexing (Chromium i7 Sample Index Plate N Set A for ATAC PN-3000427; Chromium i7 Sample Dual Index Plate TT Set A for ATAC PN-3000431). Quality control and quantification of the amplified cDNA were assessed on a Bioanalyzer (High-Sensitivity DNA Bioanalyzer kit). Sequencing was carried out at the NYGC on Illumina Novaseq using 98+26 paired-end reads.