title: GSM9231874: LUAD tumoroid scRNA-seq; Homo sapiens; RNA-Seq
library_construction_protocol: Tumoroids were isolated from Matrigel using Cell Recovery Solution (Corning), dissociated using TryLE, and FACS sorted into single cells. Single-cell RNA-seq was performed on 10x Genomics Chromium platform. The scRNA-seq libraries were prepared based on the manufacturer's protocol for 3' end reading. The individual transcriptomes of encapsulated cells were barcoded during RT step and resulting cDNA purified with DynaBeads followed by amplification per manual guidelines. Next, PCR-amplified product was fragmented, A-tailed, purified with 1.2X SPRI beads, ligated to the sequencing adapters and indexed by PCR. The indexed DNA libraries were double-size purified (0.6–0.8X) with SPRI beads and sequenced on Illumina NovaSeq S4 platform (R1 – 26 cycles, i7 – 8 cycles, R2 – 70 cycles or higher). The raw sequencing data were then demultiplexed and aligned to the mouse genome using 10X CellRanger pipeline. This generated an output matrix of mRNA counts per cell for every gene. We used the filtered matrix for all downstream analysis. For ChIP-seq, For library construction and sequencing, DNA was quantified by PicoGreen and the size was evaluated by Agilent BioAnalyzer. The KAPA HTP Library Preparation Kit (Kapa Biosystems, Cat #: KK8234) was used to prepare Illumina sequencing libraries with 0.02-5 ng of input DNA and 8-14 cycles of PCR.