Fig. 5. Axis elongation and eye defects in MAK-depleted embryos. (A) xMAK MO specifically inhibits in vitro translation of xMAK RNA. Autoradiography of in vitro translated ³⁵S-methionine-labeled proteins is shown. No suppression is observed for mMAK RNA, which lacks morpholino target sequence. (B) MAK MO inhibits xMAK translation in vivo. Western analysis with anti-Flag antibodies shows that MAK MO, but not COMO, effectively downregulated levels of Cterminally tagged xMAK in injected embryos. Loading is controlled with anti-ß-tubulin. (C) MAK MO or COMO was injected into two dorsal blastomeres of four-cell embryos. At stage 38, MAK-depleted embryos had shortened axes and eye deficiencies (see also Table 2). (D-H) Eye defects caused by MAK MO injection can be partially rescued by mouse MAK RNA in stage 38 embryos. Eight-cell embryos were injected into one animal dorsal blastomere with nßgal RNA as a lineage tracer, together with MAK MO (D,E), COMO (F) or MAK MO and mouse MAK RNA (G). (D,E) Both sides of the same injected embryo. Red staining (arrowheads) reflects lineage tracing. (H) The average eye index was calculated for each group of embryos at stage 38 as follows. 0, no visible eye; 1, severely disrupted retina with little pigmentation; 2, small or partially pigmented eye; 3, wild-type eye.