GSM7220896: Repro, WT, MEF, 2i, scRNAseq; Mus musculus; RNA-Seq
Yamanaka 4F-induced cells were trypsinized and transferred onto mitomycin-treated and SytoxBlue (Invitrogen)-stained MEF feeder cells. Cells were washed with PBS and digested with 0.05% trypsin to generate a single-cell suspension. Dead cells were stained with SytoxBlue dead cell stain and sorted by FACS. Flow cytometry-sorted cells were washed in PBS with 0.04% BSA and resuspended at a concentration of ~1000 cells/µl. Library generation for 10X Genomics v3 chemistry was performed following the Chromium Single Cell 3ʹ Reagents Kits User Guide: CG000204 Rev D. Quantiﬁcation of cDNA libraries was performed using Qubit dsDNA HS Assay Kit (Life Technologies Q32851) and high-sensitivity DNA chips (Agilent. 50674626). Quantiﬁcation of library construction was performed using KAPA qPCR (Kapa Biosystems KK4824). The library was sequenced on the Illumina NovaSeq6000 platform to target an average of ~50,000 reads per cell.