title: GSM2855485: Purified_AT2; Homo sapiens; RNA-Seq
library_construction_protocol: Lung organoid cultures from separate individuals were dissociated 4 weeks after primary plating and subjected to droplet based scRNA-seq with the 10x Genomics Gemcode Single Cell 3' platform with a 5 nucleotide UMI according to manufacturer's protocol. Cell capture, library preparation, and sequencing were performed according to manufacturer's protocol. Dissociated organoid cellular suspensions were loaded on a GemCode Single Cell Instrument (10x Genomics, Pleasanton, CA) to generate single-cell GEMs. Approximately 3-5,000 cells were loaded per channel for Lung_1 through Lung_3. Four technical replicates were generated per cell suspension, albeit for Lung_3 where one replicate failed to capture. Single-cell RNA-Seq libraries were prepared using GemCode Single Cell 3' Gel Bead and Library Kit (now sold as P/N 120230, 120231, 120232, 10x Genomics) as per Zheng et al. Sequencing libraries were loaded at 2.1pM on an Illumina NextSeq500 with 2 × 75 paired-end kits using the following read length: 98bp Read1, 14bp I7 Index, 8bp I5 Index and 5bp Read2. Note that Lung_1 through Lung_3 libraries were generated before the official launch of GemCode Single Cell 3' Gel Bead and Library Kit. Thus 5bp UMI was used (the official GemCode Single Cell 3' Gel Bead contains 10bp UMI.). For the purified AT2 library, 10,000 cells were targeted in a single technical replicate. Libraries were generated with the 10x Genomics Chromium Single Cell 3' platform v2 according to the manufacturer's protocol. The library was sequenced using paired-end sequencing (26bp Read 1 and 98bp Read 2) with a single sample index (8bp) on an Illumina NextSeq 500.