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Figure 1. Marveld3 expression during Xenopus embryogenesis and morpholinos efficiency. (A) Analysis of marveld3 transcript expression by semi-quantitative RT-PCR in non-fertilized eggs (NF) and whole embryos from stage (St) 4 to 35; gapdh expression was used as a positive control; bp: base pairs. (B) Quantification of marvelD3 expression levels. Quantitative PCR was performed with two different sets of primers for marvelD3 and a pair of primers amplifying odc as a normaliser. Shown are means ± 1 SD of measurements with three independent mRNA isolations per developmental stage. ANOVA values are provided in the graph legend. The indicated p-values in the graph were calculated with t-tests comparing to the corresponding NF values. (C) MD3A (green) and MD3B (blue) morpholino sequences are indicated in the 5′-end of the Xenopus marveld3 sequence; the start codon is indicated in red. (D) Analysis of MarvelD3 depletion by immunofluorescence was performed in animal caps derived from stage 8 embryos that had been injected with control or MD3AB morpholinos into both blastomeres at the 2-cell stage. MarvelD3 (red) and E-cadherin (adherens junction marker; green) expression was analyzed by immunofluorescence in animal caps explants. Scale bar, 100 μm; NI, non-injected embryos. (E) Ratio of fluorescence intensity at junctions for MarvelD3 and E-cadherin. Mann Whitney test p values are on the graph; the number of cells counted is indicated in brackets on the graph bars; black bar, NI animal caps; grey bar, control morpholino- and red bar, MD3AB morpholino-injected animal caps; red bar, MD3AB morpholino-injected animal caps. |
