Figure 2.
Sall1 and Nanog Reprogram EpiSCs and Influence ESC Differentiation
(A) Reprogramming efficiencies of Sall1, Nanog, and Oct4 in Oct4-GFP EpiSCs stably transfected with CRISPRa or cDNA (Oct4-GFP+ve colonies, mean of 3 independent experiments ± SD, ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001).
(B) Morphology of Oct4-GFP+ve colonies at day 20 in 2i/LIF is similar to ESC colonies. No colonies were observed in untransfected or mock transfected EpiSCs.
(C) qRT-PCR expression profiles of pluripotency markers and EpiSC markers in iPSC colonies normalized to Gapdh and relative to EpiSCs (mean of 3 independent experiments ± SD).
(D) Chimeric mouse produced with CRISPRa Sall1-induced PSCs injected into C57B/6 blastocyst.
(E) Flow cytometric analysis of Rex1-GFP+ve cells cultured in EpiSC medium at the timepoints indicated. Cells were stably transfected with Sall1 or Nanog cDNA, or empty vector and cultured in EpiSC medium (mean of 3 independent experiments ± SD, ∗∗∗∗p < 0.0001 versus PBCAG:Empty).
(F) Number of Rex1-GFP+ve ESC colonies recovered after ESCs were converted in EpiSC medium at indicated timepoints (600 cells plated at time point zero; mean of 3 independent experiments ± SD, ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001 versus PBCAG:Empty).