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Fig. 1. Xsna expression in the mesoderm and ectoderm. Distribution of Xsna mRNA shown by in situ hybridisation. Scale bar, 400 mm in
A-C and E; 200 mm in D and F. Stage numbers shown in top left (A) Stage 9, almost sagittal section. Xsna transcripts in most-vegetal tier
of marginal zone on dorsal side. bc, blastocoel; d, dorsal; v, ventral (B) Stage 11. Oblique section between horizontal and parasagittal,
with the most-dorsal side on the left. Superficial layer of marginal zone cells, suprablastoporal endoderm (se), as well as mesoderm (m),
expresses Xsna during involution through blastopore (b). Arrows indicate first appearance of transcripts in ectoderm (ec). (C) Stage 12.
Transverse section approximately one third from the anterior. Xsna expression (arrows) in both superficial (sf) and deep (dp) ectoderm
(ec). (D) Stage 18. Transverse section at the level of the head somites. Xsna transcripts are present in lateral plate mesoderm (lp) but not
prospective myotome (my). The labelled dorsolateral and ventromedial somite compartments are probably prospective dermatome (dm)
and sclerotome (s). Premigratory cephalic neural crest (c) and superficial ectoderm cells, on the tips of the neural folds (f) are also
labelled. (E) Stage 23. Transverse anterior trunk section. Absence of Xsna transcripts from notochord (n), myotome (my) and pronephros
anlage (pn) in the dorsal lateral plate, while transcripts are still detected in the dermatome (dm), sclerotome (s) and the rest of the lateral
plate (lp). Transcripts present in roofplate and trunk neural crest (rp + c) in and above the closed neural tube, respectively. (F) Stage 39.
Horizontal head section showing branchial arches (ba) with branchial arch cartilages (bc) containing Xsna transcripts. a, anterior; p,
posterior.
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Fig. 2. Organisation of the Xenopus snail gene. (A). Sequence of
5¢ region of Xsna. Sequences in bold are possible sites for Oct1,
Sp-1, B2 in TF111A promoter and AP4 (a-d) respectively (e)
Translational start of endogenous Xsna. Numbers relate to
transcriptional start (0). Lower case letters, sequence of CAT
leader. (B). Sequences at splice sites. Upper case letters, Xsna
cDNA sequence with numbers indicating position in cDNA of
bold-lettered bases.
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Fig. 3. Start site of transcription. Extension products of an
oligonucleotide complimentary to bases 77-96 of Xsna were
obtained by reverse transcription using total RNA from (a) stage
12 embryos (b) stage 20 heads (c) stage 20 posterior parts. A
DNA sequence using the same primer is shown in parallel (NB,
the sequence tracks show the complement of the Xsna promoter).
The translational start is at +33.
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Fig. 4. Effect of
promoter sequences on
expression of the CAT
reporter gene. CAT
constructs containing
the Xsna promoter are
shown (left) with black
shading in the region
affecting the
quantitative expression
most strongly.
Corresponding CAT
assays are shown on the
right. Open triangle,
labelled substrate;
closed triangle, product.
Assays were made on 4
stage 11 embryos. ID-
45/+75, internal
deletion; ui, uninjected.
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Fig. 5. Patterns of b-gal expression in embryos obtained from fertilized eggs injected with promoter constructs. Fertilised eggs were
injected with the full-length promoter (2 kb) except D and E which were injected with D -160. No significant difference was observed
between the two constructs. b, blastopore; d, dorsal midline; e, endoderm; v, ventral. (A) Stage 10.25. b-gal expression in mesoderm(m).
(B) Stage 10.5, cut through dorsal midline. Expression in mesoderm(m) and in animal cap ectoderm. (C) Stage 11. b-gal expression in
prospective neural folds (n) on lateral aspect of embryo. (D) Stage 13. b-gal expression seen in thin sections in the prospective neural
folds(n) and lateral plate mesoderm (lp). Black triangles, expression in superficial ectoderm; white triangles expression in deep ectoderm
although some superficial layer cells are also labelled. (E) Stage 13. b-gal expression in neural fold but not in neural plate (np) or ventral
ectoderm. (F) Stage 14. Transverse section through middle of embryo showing b-gal expression in lateral plate (lp) and in the left side
somite. (G) Stage 19. b-gal expression in roof of neural tube (n) and in the cephalic neural crest (cn); (bleached embryo). (H) Stage 25.
Lateral view of expression in cephalic (cn) and trunk neural crest (tn). (I) Stage 33/34. b-gal expression in somites (s) and in the branchial
arches (ba).
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Fig. 6. b-gal expression in mesoderm and ectoderm of embryos obtained from fertilized eggs injected with truncated promoter constructs.
b-gal constructs injected into embryos are on the left. Frequency of occurrence of expression in mesoderm and neural folds at stage 18 is
shown on the right. Embryos were scored as positive if any b-gal-positive cells were found in mesoderm or neural folds by dissection. ID-
45/+75, internal deletion. Bar on histogram, s.e.m.
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Fig. 7. Patterns of b-gal expression in embryos obtained from fertilized eggs injected with E-promoter constructs. Fertilised eggs were
injected at the 2-cell stage in both blastomeres with construct D -96. b, blastopore; d, dorsal midline; m, mesoderm; n, neural folds;
v, ventral. (A) Stage 11. Thin section, b-gal expression in prospective neural folds and not mesoderm. (B) Stage 11.5, expression in the
presumptive neural folds (bleached embryo). d, dorsal midline; a, anterior; p, posterior. (C) Stage 13, expression in neural folds (n).
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Fig. 8. Effect of Injecting E-promoters into specific blastomeres.
The C1, A2, and A4 blastomeres at the 32-cell stage (Dale and
Slack,1987) and at the 1-cell stage were injected with either the Epromoter
(D -96) or the complete short promoter (D -160).
*Occurrence of b-gal-stained cells in neural folds, epidermis or
mesoderm were scored as in Fig. 6.
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Fig. 9. Effect of mesoderm-inducing factors and the vegetal pole
cells on expression of promoter-CAT fusions. Animal cap
explants obtained from embryos injected with the full-length WTCAT
construct were cultured in media containing growth factors
or placed in a conjugate with vegetal pole cells. CAT activity was
measured in the explants at the equivalent of stage 10.5. Caps (30-
45) were homogenised and the equivalent of 4 caps were used per
assay. (A) Embryos at stage 10.5. (B) Caps alone. (C) Caps +
bFGF 9 u per ml. (D) Caps + activin 20 u per ml. (E) Caps +
bFGF and activin. (F) Caps + vegetal pole. (G) Vegetal pole
alone. Acetylated product is at the top of the figure.
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