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Fig. 1. Early and late shifting nuclear antigens in
embryos of X. laevis. Embryos at late blastula stage 9
(A,B) and gastrula stage 12 (C,D) were fixed with
formaldehyde and embedded with wax. Sections were
stained with mABs b2-2B10 (A), and 32-5B6 (B,C). The
same section shown in C was counterstained with DAPI
to locate all cell nuclei (D). Bar 100um.
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Fig. 2. Nucleocytoplasmic distribution of nucleoplasmin in embryos of X. laevis. Embryos after the 7th (A,B), 11th
(C,D), and about the 13th cleavage division (E, F) were fixed according to Romeis and embedded in wax. Sections were
stained with mAB b7-lA9 that binds to nucleoplasmin. Details of the sections shown in the left panels (A,C,E) are
shown in the right panels (B,D,F). Bar 100um.
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Fig. 3. Kinetics of pronucleus formation after injection of sperm nuclei into unfertilized eggs of X. laevis. Sperm nuclei
were isolated and injected into eggs as detailed in Materials and methods. Eggs were fixed with TCA 15min (A,B),
30min (C,D), and 60min after injected, embedded in wax, and sectioned. Sections were stained with mAB b2-2B10,
which binds to N, (A,C,E) and counterstaind with DAPI (B,D,F). Nuclei that do not swell do not take up the antigen
to a significant degree (E,F). Bar 100um.
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Fig. 4. Multiple pronuclei formed after injection of sperm nuclei into unfertilized eggs. X. laevis eggs were fixed with
TCA 2h after injection of sperm nuclei. Wax sections were stained with mABs b2-2B10 (A) and L6-8A7, which binds to
lamin Lm (B). Counterstaining with DAPI showed that all nuclei had taken up both antigens (not shown). Bar 100um.
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Fig. 5. Selective uptake of early-shifting nuclear antigens
into injected sperm nuclei. Eggs were fixed with TCA 2h
after injection of sperm nuclei. Wax sections were stained
with mABs 32-4A1 (A), 37-1B2 (C), P3 control antibody
IgGl secreted by mouse cell line P3x63 Ag8 (E).
Counterstaining of each of these sections with DAPI is
shown on the right panels (B,D,F). Bar 50um.
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Fig. 6. Selective uptake of early-shifting antigens into injected erythrocyte nuclei. Eggs of X. laevis were fixed with TCA
2h after injection of erythrocyte nuclei. Wax sections were stained with mABs L6-8A7 (A), b2-2B10 (C), and 37-1B2
(E). Counterstaining of the same sections with DAPI is shown in the right panels (B,D,F). Bar 50m.
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Fig. 7. Artificial pronuclei formed after injection of A-DNA. Unfertilized eggs of X. borealis were injected with 5-10ng
of A-DNA each. 2h after injection, eggs were fixed with TCA. Wax sections were stained with mABs b2-2B10 (A) and
32-5B6 (C). Counterstaining of each section with DAPI is shown on the right panels (B,D). Bar 50um.
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Fig. 8. Nucleocytoplasmic distribution of oocyte proteins synthesized de novo. Oocytes of X. laevis were labelled with
[35S]methionine for 5 h (A,B) or 15 h (C,D), manually enucleated and protein preparations of either four total oocytes
(A,C) or of 30 nuclei were subjected to two-dimensional gel analysis and fluorography as detailed in Materials and
methods. After fluorography, the areas at the position of the prominent nuclear and cytoplasmic proteins indicated in
the figure were excised from each of the four gels and the amount of radioactivity in each area determined by liquid
scintillation counting. The results are given in Table 2. A, actin; T, /J-tubulin; N,, antigen b2-2B10; 5, antigen 35-1A7;
7, antigen 37-1A9; 21, antigen 32-5B6 (Dreyer & Hausen, 1983; Dreyer et al. 1985). Nucleoplasmin is not labelled under
these conditions.
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Fig. 9. Phosphorylation of germinal vesicle proteins in
oocytes and maturing eggs of X. laevis. Oocytes in the
absence (A,B) or presence (C) of progesterone were labelled
with [32P]phosphate for 4h as detailed in Materials and
methods. Protein preparations were subjected to twodimensional
gel analysis. The gels were stained with silver
and analysed by autoradiography. (A) Oocyte proteins
stained with silver. Isoelectric focusing was from right to left,
SDS-polyacrylamide gel electrophoresis from top to bottom.
(B) Autoradiograph of the gel shown in A. (C)
Autoradiograph of gel showing 32P-labelled proteins from
maturing egg. Protein patterns of oocytes and of maturing
eggs were not significantly different after silver staining (not
shown). Germinal vesicle proteins are numbered as
previously. Nu, nucleoplasmin.
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Fig. 10. Cell cycle arrest with cycloheximide. Embryos of
X. laevis were transferred to lOO^gml"' of cycloheximide
in MBSH at the 64-cell stage and incubated therein for
4h, when controls had reached stage 9+. Experimental
and control embryos were fixed by freeze substitution,
and wax sections were stained with mAB 32-5B6 (A,C)
and b2-2B10 (B,D). (A,B) Controls; (C,D) embryos
treated with cycloheximide. Bar 100um.
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