GSM3081854: WT 48h; Mus musculus; RNA-Seq
NSCs were resuspended at 103 cells/µL in PBS containing 0.04% (w/v) BSA to minimize cell loss and aggregation. ~17,500 cells were partitioned into nanoliter-scale Gel Bead-In-EMulsions (GEMs) to achieve single cell resolution for a maximum of 10,000 individual cells per sample. Utilizing the v2 Chromium™ Single Cell 3' Library Kit and Chromium instrument (10x Genomics), poly-adenylated mRNA from an individual cell was tagged with a unique 16 basepair 10x barcode and 10 basepair Unique Molecular Identifier. Full length cDNA was amplified to generate sufficient mass for library construction.  Enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size (~400 basepair) for the library. The final library was sequence-ready and contained four unique sample indexes. The concentration of the 10x single cell library was determined through qPCR (Kapa Biosystems). The libraries were normalized, pooled, and sequenced with a custom recipe (26-8-98) on the HiSeq4000 platform (Illumina). Five single cell libraries were sequenced across an entire HiSeq4000 flow cell targeting ~45,000 reads per cell.
GEO Accession
GSM3081854