GSM4253871: Organoid Ngn3-RFP positive single cell transcriptomes, ISX-9 treated; Mus musculus; RNA-Seq
Samples were processed by Single Cell Discoveries B.V. according the SORT-seq method (Muraro et al., 2016). Ngn3-RFP-positive cells were sorted into 384-well plates with 5 μl of vapor-lock (QIAGEN) containing 100-200 nl of RT primers (consisting of a 24 bp polyT stretch, a 4bp random molecular barcode (UMI), a cell-specific 8bp barcode, the 5′ Illumina TruSeq small RNA kit adaptor and a T7 promoter), dNTPs and and mRNA ERCC Spike-Ins (Agilent). Plate was spun and frozen at −80°C. Cells were then lysed by 5 min at 65°C. mRNA of each cell was reverse transcribed, converted to double-stranded cDNA, pooled and in vitro transcribed for linear following the CEL-Seq 2 protocol (Hashimshony et al., 2016). Illumina sequencing libraries were then prepared with the TruSeq small RNA primers (Illumina) and sequenced paired-end at 75 bp read length the Illumina NextSeq.
GEO Accession
GSM4253871