GSM4453843
GSM4453843: Day14 Ventral cDNA; Homo sapiens; RNA-Seq
MiSTR tissue was divided into two halves along the gradient, one side used to collect the five A-E samples for mRNA analysis. The other side was dissociated into a single-cell suspension using Accutase (Thermofisher), for day 0, day 1 and day 2 samples, or with Neural dissociation kit (P) on a GentleMACS dissociator (both from Miltenyi Biotec) for day 14 samples. The cells were then frozen in CS10 CryoStor (Stemcell Technologies) and stored at -150oC until needed. In the meantime, each sample underwent mRNA analysis for quality control selection for scRNA-seq. Cryopreserved cells, quality control cleared, were thawed, counted and mixed in equal numbers into a final single-cell suspension of 350 to 500 cells/ml solution in 0.01% BSA in PBS. For 0 to 48h tissues, cells from 3 biological MiSTR tissue replicates were pooled prior to sequencing. For day 14 tissues, the sequenced samples contained a mix of cells from regions A to E (n=1-3 biological replicates per region). A total of 9000-17000 cells were loaded per lane onto the 10X Chromium (10X Genomics) . For day14 ventral Cells from region A-E were hashed with hashtagging antibodies and then loaded on 10X chromium to generate hashing and cDNA libraries simultaneously. Chromium Single Cell 3' Reagent Kits (v2 Chemistry).
GEO
GSM4453843
GEO Accession
GSM4453843