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Fig. 1 | Stem Cell Research & Therapy

Fig. 1

From: Targeted reversion of induced pluripotent stem cells from patients with human cleidocranial dysplasia improves bone regeneration in a rat calvarial bone defect model

Fig. 1

Generation of cleidocranial dysplasia (CCD) patient-specific iPSCs and genome editing. a Identification of heterozygous runt-related transcription factor 2 (RUNX2) mutations in each CCD. b Functional domains of RUNX2 proteins and RUNX2 exon organization. c Morphology of established CCD patient-derived induced pluripotent stem cells (CCD-iPS). d–f Confirmation of the pluripotency of the CCD-iPSCs. d RT-PCR analysis of ESC marker genes. e RT-PCR analyses of various differentiation markers for the three germ layers. Target genes included AFP, FOXA2, and SOX17 (endoderm), T and MSX1 (mesoderm), and MAPs (ectoderm). β-actin was used as an internal control. f Teratoma formation and histology. Teratoma contained tissues of three embryonic germ layers: cartilage (mesoderm), gut-like epithelium tissues (endoderm), and neural tube-like structures (ectoderm). g Karyotype analysis (Q-band) of CCD1- and CCD2-iPSCs. h Clustered regularly interspaced short palindromic repeats (CRISPR)/single-guide RNA (sgRNA) targeting of the RUNX2 gene on chromosome 6. i CRISPR-mediated genome editing of CCD1-iPSCs. j Confirmation of the RUNX2 gene correction by sequence analysis. k Confirmation of the pluripotency of the Reverted iPSCs (Rev1-iPSCs). l Karyotype analysis (Q-band) of Rev1-iPSCs. Abbreviations: D, differentiated; U, undifferentiated; Q/A, glutamine/alanine-rich domain; RHD, runt homology domain; NLS, nuclear localization signal; NMTS, nuclear matrix-targeting signal; FRT, flippase recognition target; PGK, phosphoglycerate kinase

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