GSM8727809: DKOC; Mus musculus; RNA-Seq
Mouse organoids were dissociated into single cells and resuspended in organoid culture medium for the library preparation. The Chromium Next GEM Single-Cell 3ʹ Reagent Kit (10× Genomics) was used according to the manufacturer's protocols. Briefly, organoid cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscribed to cDNA, which contains an Illumina R1 primer sequence, Unique Molecular Identifier (UMI), and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR, and the appropriate sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction, Illumina R2 primer sequence, paired-end constructs with P5 and P7 sequences, and a sample index were added.