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Fig. 6. gli2 acts through a transcriptional activation mechanism and affects the expression of several NC markers. (A–B) Gli-Luciferase reporter assay. (A) Schematic representation of the Luciferase assay experiment. (B) Transcriptional activity of Gli2 in NC explants. Intensities are shown as relative units of Luciferase normalized to the internal control Renilla Luciferase. An increase in Gli2 and Ihh function increases the expression of the Gli-dependent reporter construct. (C-D) RT-qPCR analysis of Gli2 function on the expression of NPB and NC markers. Each color coded bar represents the relative expression level of the markers for each condition (gain and loss-of-function of gli2). All the expression levels denote Ct values that were normalized first to the expression level of gadph, and then they were normalized to the expression of each gene in the uninjected wildtype condition, and are presented as fold change. (C) Schematic representation of the grafting experiment. At stage 15–16 total RNA was isolated and then RT-qPCR was performed. (D) Expression levels of the markers pax3, hairy2b, snail1, snail2, twist-b, e-cad and n-cad. Data are presented as mean ± s.d. of triplicates of two independent qPCR experiments. Reference: **, P < 0.05; ***, P < 0.001 (P-values correspond to Student's t-Test). Image published in: Cerrizuela S et al. (2018) Copyright © 2018. Image reproduced with permission of the Publisher, Elsevier B. V. Larger Image Printer Friendly View

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