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Using RT-PCR and in situ hybridisation, we have analysed the temporal and spatial expression patterns of Xenopus Fox genes of subclass N. By screening cDNA libraries and by RT-PCR using embryonic RNA and primers derived from EST analyses, we could isolate FoxN2, FoxN4, FoxN5 and different isoforms of FoxN3. FoxN2 and FoxN3 transcripts were found during all developmental stages including early cleavage and tailbud stages. FoxN5 transcripts were only present at early cleavage stages, while FoxN4 expression began after midblastula transition. Spatial expression of FoxN2 was first detected in the early eye field and later, in the branchial arches, the vagal ganglion and in the developing retina. FoxN3 transcripts were found within the animal cap. In post-gastrula embryos, neural crest cells and the early eye field showed strong expression of FoxN3. At late tadpole stages, the branchial arches were stained. FoxN4 was expressed in the early eye field and later in the developing retina cells, the nephrostomes of the pronephric kidney and in the midbrain. A ubiquitous expression of FoxN5 was found in early cleavage stage embryos.
Fig. 4 (Left). Whole mount in situ hybridisation of FoxN2. (A) Stage 26, lateral view; (B) magnification of the head of a stage 33/34 embryo, lateral view; (C) lateral view of a stage 29 embryo. The red lines in (A-C) denote transversal sections shown in (D-F). (D) Stage 26; (E) stage 29; (F) stage 33/34. (G) A transversal section through the head of an embryo at stage 36. ba, branchial arches; l, lens; prl, presumptive retinal layer; rh, rhombencephalon; r, retina; vg, vagal ganglion. The fat arrow in (E) denotes the FoxN2 stained cell layer, lateral to the ventricular zone.
Fig. 5 (Above). Whole mount in situ hybridisation of FoxN3. (A)
Transversal razor blade sectioned 8-cell stage embryo, animal cells are on top. (B) Dorsal view of a stage 23 embryo. Lateral views of (C) stage 26, (D) stage 29 and (E) stage 38 embryos. (F) Transversal section of an embryo at stage 38. The red line in (E) indicates plane of section shown in (F). ba, branchial arches; hm, head mesenchyme; l, lens; vg, vagal ganglion.
Fig. 6. Whole mount in situ hybridisation of FoxN4. Anterior view of a (A) stage 18, (B) stage 21 and (C) stage 23 embryo. (D) Half-lateral view of a stage 26 embryo. Lateral view of a (E) stage 29/30 and (F) stage 33/ 34 embryo. The red line in (F) denotes the plane of the transversal sections shown in (H-I). (G) Transversal section of the head of a stage 26 embryo. (H) Section of a stage 33/34 embryo. (I) Section of a stage 38 embryo. cmz, ciliary marginal zone; l, lens; mb, midbrain; n, nephros- tomes; prl, presumptive retinal layer; r, retina.