GSM7770156: Differentiation Ctrl; Homo sapiens; RNA-Seq
Cells were lysed by 5 min at 65 ℃, after which RT and second strand mixes were dispersed with the Nanodrop II liquid handling platform (GC biotech). Aqueous phase was separated from the oil phase after pooling all cells in one library, followed by IVT transcription. The CEL-Seq2 protocol was used for library prep. Primers consisted of a 24 bp polyT stretch, a 4bp random molecular barcode (UMI), a cell-specific 8bp barcode, the 50 Illumina TruSeq small RNA kit adaptor and a T7 promoter. mRNA of each cell was then 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