GSM8138121: Melanocyte Stem Cells (Sample 3); Mus musculus; RNA-Seq
Flow Sorted Kit+/Cd45- (McSCS) and single cell suspension for dermis sample Cell suspension, 10x barcoded gel beads, and oil (all were included in 10xGenomics Chromium Next GEM Single Cell 3' Kit v3.1 PN 1000268)were loaded into 10x Chromium™ Single Cell Chip to capture single cells into nanoliter-scale oil droplets by 10xChromium™ Controller and generate Gel Bead-In-EMulsions (GEMs). cDNA from single cells were synthesized and barcoded by incubation of the GEMs in a thermocycler machine, and then purified from GEMs by DynaBeads (10xGenomics PN 2000048). cDNA was pre-amplified by PCR to generate sufficient mass for sequencing library construction. The single cell cDNA libraries were constructed by following 10xGenomics 3' NextGem 3.1 version kit instruction, briefly by following steps of cDNA fragmentation, end repair & A-tailing, and size selection by SPRIselect beads (Beckman Coulter, PN B23318),adaptor ligation, and sample index PCR amplification, and then SPRIselect beads size selection again. The final constructed 3'-biased single cell libraries were sequenced by Illumina Nextseq500 machine, targeting 20,000 read pairs/cell. and the sequencing cycles consisted of 28bp for read 1, 91bp for read 2 and 10bp for i7 and i5