GSM7912211: Flow-Sorted e15.5 endocrine-committed and endocrine-progenitor cells-DKO 1; Mus musculus; RNA-Seq
Each clean single cell suspension was counted with hemocytometer under microscope for cell number and cell viability. The single cell suspension had a viability of >90%. Approximately 200-1000 targeted cell recovery per sample was applied to a single cell master mix with lysis buffer and reverse transcription reagents, following the Chromium NextGEM Single Cell 3' Reagent Kits User Guide, CG000204 Rev D (10X Genomics, Inc). Along with the single cell gel beads and partitioning oil, the single cell master mixture containing the single cell suspension was dispensed onto a Single Cell Chip G in separate wells, and the chip loaded to the Chromium Controller for GEM generation and barcoding, followed by cDNA synthesis and library preparation. At each step, the quality of cDNA and library were examined by Bioanalyzer and Qubit. The resulting barcoded libraries were pooled in equal molarity and sequenced in a custom program for 28 bp plus 91 bp paired-end sequencing on Illumina NovaSeq 6000 sequencer (Illumina, Inc.). Approximately 50000 reads per cell were generated and 91% of the sequencing reads reached Q30 (99.9% base call accuracy). A Phred quality score (Q score) was used to measure the quality of sequencing.