XB-IMG-81395
Xenbase Image ID: 81395
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Fig. 3. The effect of RA on primary neurogenesis requires xEmGCNF. (A)
Schematic representation of the experimental system. Embryos were
injected at the two-cell stage into each blastomere. Animal caps were
explanted at stage 8.5 and cultivated until stage 17 equivalent. Total RNA
was extracted and semi-quantitative RT-PCR was performed. (B) Knockdown
of xEmGCNF abolishes the increasing effect of RA signaling on the
expression of primary neurogenesis markers. Injection of 40 pg noggin
mRNA directed the ectoderm explants to an anterior neural fate (Nogg,
lanes 2). Further posterior specification of the neuroectoderm by an
enhanced RA signaling (RA, lanes 3) was achieved by coinjection of
mRNAs encoding the retinoid receptors xRARa2 (250 pg) and xRXRb
(250 pg) and treatment of the neuralized explants with 1027 M RA. In
addition, embryos were co-injected with 42 ng of the STA (lanes 1) or
42 ng of the MOR (lanes 4). Expression of markers of neuronal end
differentiation (NST; XDelta-1; Xngnr1 and NeuroD), pan-neural markers
(N-CAM and Nrp1) was monitored by RT-PCR using gene-specific
primers. Loading control ODC RT, negative control ODC 2 RT, whole
embryo WE. (C) Whole mount in situ hybridization of stage 18 embryos
to detect NST (C,D) and XDelta-1 mRNA (E and F) after co-injection of
42 ng of asMOR and 150 pg of b-galactosidase mRNA into one cell at the
two-cell stage. b-Galactosidase staining in red identifies the injected
hemispheres to the right. Anterior is at the top of each figure. D and F show
details of C and E, respectively. Image published in: Barreto G et al. (2003) Copyright © 2003. Image reproduced with permission of the Publisher, Elsevier B. V.
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