Custom Taqman probes were ordered for Nanog, Otx2, Ascl2, and beta-actin (Thermo Fisher Scientific). Single cells were collected directly into 96-well plates by flow cytometry (MoFlo, Beckman Coulter) and reverse transcription was performed as described previously (45 (link)). Cells that had Ct value of beta-actin >14 and no amplification were excluded from the analysis.
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