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Fig. 1. Genomic organization and expression of the Tsg gene. (A) Schematic
representation of the wild-type and targeted alleles of Tsg. The
5' probe used for the Southern blot is indicated (probe). The
oligonucleotides used for PCR genotyping are shown as arrows: Tsg22, Tsg23 and
lacZ3. The position of the exons is indicated by black boxes and the
recombination arms shown in red. The following restriction sites are
indicated: B, BamHI; K, KpnI; N, NsiI; S,
SacI. (B) Southern blot analysis of the 5' genomic region of
the Tsg gene. DNAs extracted from wild-type, heterozygous and
homozygous Tsg mutant animals were digested with SacI. The
170 bp SacI-NsiI probe fragment detected a wild-type band of
9.2 kb and a knock out (KO) band of 3.7 kb. (C) PCR genotyping of mice
carrying the Tsg mutation. The wild-type band of 329 bp was obtained
using the Tsg22 and Tsg23 primers. The KO band of 480 bp was obtained using
the Tsg22 and lacZ3 primers. (D) RT-PCR analysis of RNAs extracted
from E13.5 wild-type and Tsg-/- embryos showing the
absence of Tsg mRNA in Tsg-/- embryos.
HPRT expression was used as a control. (E-H,E'-H')
Comparison of the expression pattern of Tsg by whole-mount in situ
hybridization and ß-galactosidase staining. (E) Tsg is expressed
in the anterior visceral endoderm and the primitive streak. (F,G) As
gastrulation proceeds, Tsg is found in the mesodermal layer. (H) At
E8.5 Tsg is present in the endoderm of the gut, the ventral neural
tube and the dorsal region of the eye vesicle. (E'-H') The
expression of the lacZ transgene recapitulates the expression of
endogenous Tsg. (I) Transverse section through the anterior region of
an E8.0 embryo showing Tsg mRNA in the endoderm of the foregut,
ventral neuroectoderm and the head mesenchyme. (J) Section of an E8.5 embryo
showing the expression of Tsg mRNA in the basal diencephalon. ave,
anterior visceral endoderm; bd, basal diencephalon; bm, basal mesencephalon;
ev, eye vesicle; en, endoderm; fg, foregut; hg, hindgut; hm, head mesenchyme;
mw, mesodermal wings; ne, neuroectoderm; ps, primitive streak; A, anterior; L,
left; P, posterior; R, right.
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