Knockdown of lpp disrupts convergence and extension movements. (A, B) Validation of the morpholinos: (A) quantitative RT-PCR using total cDNA from tailbud stage embryos as a template, shows an 80% reduction of lpp expression after injection of Lpp-MO/3e3i but not after injection of Lpp-MM/3e3i; (B) Lpp protein levels of prim-5 stage embryos are strongly reduced after injection of Lpp-MO/ATG but not after injection of Lpp-MM/ATG, as shown by quantification of the intensities of the protein bands after Western blot analysis (cumulative results of three experiments). Error bars represent standard errors of the mean. (C–L) Morpholinos (Lpp-MO/ATG and Lpp-MO/3e3i) or mismatch control morpholinos (Lpp-MM/ATG and Lpp-MM/3e3i) were injected at the one- to two-cell stage and morphology was assessed at different stages. (C–H) tailbud stage, lateral view, dorsal to the right; the anterior-most structure is indicated with an arrow. (I, J) 5-somite stage, lateral view, dorsal to the right. (I′, J′) 5-somite stage, dorsal view; drawings schematically show the notochord and somites of the embryos in the pictures. (K, L) prim-5 stage, lateral view, anterior to the left. Abbreviations: NI, non-injected; ML, mediolateral axis; AP, anterior–posterior axis. (M) Zebrafish embryos were injected with Lpp-MO/3e3i (5 ng) or synthetic mRNA (7.5 pg) encoding full-length Lpp, or were co-injected with Lpp-MO/3e3i (5 ng) and synthetic lpp mRNA (7.5 pg). At tailbud stage, the embryos were scored morphologically for the presence of C&E defects, as depicted in panels E and H. The phenotypes of the embryos of three independent experiments were scored and the percentages of normal embryos, embryos with C&E defects (“CE defects”) and embryos with defects other than C&E defects (“other defects”) are indicated. n, total number of embryos injected for each condition.
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