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Fig. 7. Expression of somitic, lateral plate, blood and endothelial marker genes in Pax8 morphants. (A) Expression of somitic (myoD and mlc) and lateral plate markers (foxf1). Embryos were injected with MoPax8 at the 4-cell stage, in the equatorial region of both left blastomeres. They were cultured until tailbud stage 25 (a, a, b, b) or neurula stage 18 (c, c), fixed and analyzed by whole-mount in situ hybridization with the indicated probes. For each embryo, injected (a, b, c) and uninjected (a, b, c) sides are shown. Interference with Pax8 function does not modify the expression pattern of neither myoD, mlc and foxf1. (B) Expression of blood (scl and lmo2) and endothelial markers (msr and flk1). Embryos were injected with MoPax8 (dg) or MoC (hk) at the 4-cell stage, in the equatorial region of both left blastomeres. They were cultured until tailbud stage 27 and processed for whole-mount in situ hybridization with the indicated probes. For each embryo, injected (dj, hk) and uninjected (dg, hk) sides are shown. Double in situs with pax8 and lmo2 probes are shown in whole mount (d, d) as well as in section performed at the level of the anterior (l) and posterior (m) part of the pronephros anlage and in the posterior region of the embryo (n). The dotted line delineates injected (left) and uninjected (right) halves. In Pax8 morphants, the expression domain of scl, lmo2, msr and flk1 is enlarged in the anterior region where the tubule anlage normally forms. Double in situ hybridization with pax8 and lmo2 probes shows a much stronger lmo2 signal dorsal to pax8 expression domain on the injected side at the level of the pronephros anlage (l, m). In A and B, pax8 is revealed in light blue while the other genes are revealed in dark blue. Scale bar: 200 m, (C) Expression of lmo2 in Pax8 morphants in the absence of cell division. Embryos were injected with MoPax8 at the 4-cell stage, in the equatorial region of both left blastomeres. Half of them were cultivated in the presence of HUA from neurula stage 18 onwards. They were processed for whole-mount in situ hybridization at stage 27. The enlargement of the lmo2 expression domain in the anterior region in response to the loss of Pax8 (o) is maintained in HUA treated embryos (p). |
