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Additional Supporting Information may be found in the online version of this article.

FilenameFormatSizeDescription
sc-12-0552_sm_SupplFigure1.tif943KFigure S1. A) Gene expression levels of Pax3, Pax7 and Myf5 in inducible Ires-GFP day 5 control EBs. qRT-PCR was performed using the indicated TaqMan probes and the results were normalized to Gapdh endogenous levels. B) Paraxial mesoderm and myogenic marker are up-regulated by Pax3 induction in day 5 EBs. qRT-PCR was performed using the indicated TaqMan probes and the results were normalized to Gapdh endogenous levels. The graphs report the mean +SD for at least 3 independent experiments. C) Frequency of PDGFRα+FLK-1+ (P+F+), PDGFRα+FLK-1- (P+), PDGFRα-FLK-1+ (F+) and PDGFRα-FLK-1- (NEG) fractions before and after fluorescence activated cell sorting (FACS). Data are mean ±SD of 3 independent experiments.
sc-12-0552_sm_SupplFigure2.tif2368KFigure S2. Effect of Pax3 on MyoD and Myogenin expression levels at day 15 of differentiation. A) Schematic diagram showing the timing of Pax3 induction. At day 5, PDGFRα+FLK-1- cells were sorted and cultured until day 15. B-C) Western blot (B) and gene expression (C) analyses (n=2) for MyoD, MyoG and Pax3.
sc-12-0552_sm_SupplFigure3.tif1426KFigure S3. Alignment of the C-terminal region of PAX3 and PAX7 sequences from different organisms, adapted from the Conserved Domain Database (NCBI). The region in red represents the pfam12360.
sc-12-0552_sm_SupplFigure4.tif617KFigure S4. Gene expression analysis of Pax3 cell lines. RNAs were extracted from total EBs (day 5) using Trizol and used for cDNA synthesis. qRT-PCR was performed using the indicated TaqMan probes and the results were normalized to Gapdh endogenous levels and reported as relative expression to the non-induced sample. The graphs report the mean +SD for at least 3 independent experiments.
sc-12-0552_sm_SupplFigure5.pdf115KFigure S5. Pax3 represses cardiac terminal differentiation. Unsorted day 5 EBs noninduced or dox-induced from day 2 were trypsinized and plated as monolayer and cultured until day 10. A) Cells were fixed and analyzed by immunofluorescence staining with an anti-cTNI antibody, or B) harvested, lysed with RIPA buffer and analyzed with western blot using anti-MYOD, anti-cTNI and anti-GAPDH antibodies. Pax3 (lanes 1 and 6); Pax3.8 (lanes 2 and 7); Pax3 .PD-C (lanes 3 and 8); Pax3 .HD-C (lanes 4 and 9); Pax3 .PD (lanes 5 and 10). Bar: 50μm.
sc-12-0552_sm_SupplTable1.pdf27KSupplementary Table 1.

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