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Gene/Clone	Species	Stage	Anatomy Item	Experimenter
egr2	xenopus

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Experiment details for egr2

De Marco N et al. (2011) Assay

Eukaryotic initiation factor 6 (eif6) overexpression affects eye development in Xenopus laevis.

Gene	Clone	Species	Stages	Anatomy
egr2.L		laevis	NF stage 19	presumptive rhombomere

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Fig. 2. eif6 overexpression specifically inhibits eye development. (a)–(j) Embryos were injected with eif6 (400 pg) and βgal (400 pg) or GFP (400 pg) alone into one blastomere of the two-cell stage and harvested at the neurula stage. Whole mount in situ hybridisations were performed for rax (a), (b), pax6 (c), (d), otx2 (e), (f), six3 (g), (h) and sox2 (i), (j), fgf8 (k), (l), en2 (m), (n), egr2 (o), (p).GFP overexpression did not affect the expression of rax (a), pax6 (c), otx2 (e), six3 (g), sox2 (i), fgf8 (k), en2 (m), egr2 (o). Inset in (a) shows diffusion of GFP fluorescence in the whole injected side (i.s.) of a neurula. eif6 overexpression specifically reduced rax (b), pax6 (d), otx2 (f) and six3 (h) expression within the presumptive eye field (arrowheads), while neural expression of the same markers was unaffected (arrows). (j) No expression difference was found for the specific neural markers sox2, fgf8 , en2, egr2 between the eif6-injected and un-injected side.

Gene	Clone	Species	Stages	Anatomy
egr2.L		laevis	NF stage 35 and 36	forebrain , telencephalon , diencephalon

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Fig. 1. Effects of eif6 overexpression on eye formation. (a)–(f) Three different embryos injected with 400 pg eif6 mRNA and 300 pg GFP mRNA into one blastomere of the two-cell stage and harvested at stage 35. Micrographs of the two sides (a and b; c and d; e and f) of the same embryos are shown; (a), (c), and (e) are the injected sides. The eif6 overexpressors showed a range of the defective eye phenotype (arrowheads in a, c, e). The injected sides were marked by GFP fluorescence (inset in a, c, e). (g) GFP-injected side (fluorescence in the inset) and un-injected side (h) of the same embryo: overexpression of GFP alone did not affect eye development. (i)–(j) Micrographs of the two sides of the eif6-injected embryo harvested at stage 42. The eye of the injected side marked by GFP fluorescence (inset in i) recovers normal morphology. (k)–(p) Cryostat sections of stage 35 eif6-injected embryos showed a reduced eye in the injected side. Magnification of reduced eye in insets of figures k–n. (k) DAPI staining. Dasher lines border the entire thickness of neural retinas. The eye marker pax6 (l) was present in both injected and un-injected eye. However the retina layer markers showed a difference between injected and un-injected eye. Indeed, in the small injected eye otx2 (m) was diffuse and not localised in the central retina as in the un-injected eye. rbpms, (n) becomes evident only in the un-injected side. The encephalon structures were morphologically normal, forebrain (o) and midbrain (l) were labelled with pax6, hindbrain (p) was marked by egr2. (q)–(t) Cryostat sections of stage 42 eif6-injected embryos. (q) DAPI staining shows a normal stratification of retina in the injected eye compared to the control eye. (r)–(t) In situ hybridisation with rbpms, nrl, an outer nuclear layer marker and otx2, showed no difference between injected and un-injected eye, indicating that the recovered retina is structurally similar to the control retina. i.s :eif6-injected side.