Murine iPSCs were transduced with a construct: MiDR-TCRα-2A-TCRβ-2A-FoxP3 containing genes of OVA-specific TCR and FoxP3. The gene-transduced cells (DsRed+) were co-cultured on OP9-DL1-DL4-I-Ab cells in the presence of mFlt3L and mIL-7. (A) Morphology of Tregs cell differentiation on day 0, 7, 14 and 22. (B) Flow cytometric analysis for the protein expression of iPSC-derived cells on day 28. CD3+ TCRVβ5+ cells were gated as indicated (R1) and analyzed for the expression of CD4 and CD8, with CD25, CD127, CTLA-4, and FoxP3 shown for cells gated as CD4+CD8- cells (R2) (dark lines; shaded areas indicate isotype controls).