Bicc1 regulates dand5 mRNA through distinct regions of the Bicc1RE.A Quantification of right-sided pitx2 induction by co-injecting a low, ineffective m-tpMO dosage with single allele-specific bicc1 SBMO. Controls (co), m-tpMO (S or L, low), or allele (S or L) specific bicc1 SBMO alone showed wt pitx2 asymmetry. B Co-injecting m-tpMO (low) with bicc1 TPMO (L or S) impacted dand5 mRNA stability. Treatment with low concentrations of m-tpMO, single allele-specific bicc1 TPMO, and uninjected co showed wt dand5 expression at post-flow stages. C Quantification of dand5 expression. D Quantification of pitx2 asymmetry. Only in combination both suboptimal dosages of d-tpMO (low) or single allele-specific bicc1 SBMO (S or L) prevented left pitx2 expression. Wt expression was found in controls (co) and in embryos that were left-sided injected with one MO alone. MO pmol/embryo: bicc1 SBMO (L or S, 1); m-tpMO low (L or S, 0.4); d-tpMO low (L or S, 0.5). Asterisks in B mark injected side. Numbers (n) in A, C, and D represent analyzed specimens from more than independent experiments. Statistical analyses were done with one-sided Pearson’s chi-square test, which was adjusted for multiple comparisons by Bonferroni (A, D) or Bonferroni–Holm (C). n.s., not significant p > 0.05; ***, very highly significant p < 0.001. p values and listing of individual experiments can be found in the source data file. Scale bar in B represents 100 µm. st. stage, a anterior, l left, r right, p posterior.
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