scRNA-seq TF-driven forward programmed hiPSCs to multiple fates

This dataset consists of single-cell RNA-seq (Smart-seq2) data from 95 forward programmed cells derived from human induced pluripotent stem cells (hiPSCs). Cells conditionally expressing either ATOH1, ETV2 or NKX3-1 under the control of a tetracycline-response element (Tet-ON system) were co-cultured on Matrigel in 2 dimensions. Data was used to explore the forward-programming potential of these transcription factors (TFs) when cell lines with the capacity to differentiate in isolation are cultured together. Overall design: Three hiPSCs line were engineered using the PiggyBac transposition system. Vectors containing a cassette encoding one of three TFs (ATOH1, ETV2 or NKX3-1) and the gene for puromycin resistance, both under the control of the tetracycline response element (Tet-ON system) and flanked by sequences to be recognized by the PiggyBac transposase, were co-delivered with a vector encoding the PiggyBac transposase to hiPSCs by nucleofection. After antibiotic selection, cells from the three lines were mixed at a 1:1:1 ratio in mTeSR1 supplemented with the Y-27632 ROCK inhibitor and doxycycline and seeded on matrigel-coated 12-well plates. mTeSR1 medium supplemented with doxycycline was replaced every day until the end of the experiment. Four days (96 hours) after seeding, cells were dissociated and immunofluorescently labeled with fluorophore-conjugated antibodies against NCAM (CD56; neuronal marker), Vimentin (fibroblast marker) and VE-Cadherin (endothelial marker) in a standard buffer for flow cytometry.Individual cells were then sorted in a 96-well plate containing a lysis buffer for RNA isolation. Single cell cDNA libraries were prepared from each cell following the Smart-seq2 workflow. 95 fastq files are provided corresponding to each individual sorted cell; a well was left empty to be used as a negative control for the presence of RNA and cDNA.