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Fig. 2. Targeted replacement of wild-type Sox10 by mutant
Sox10 sequences in mice. (A) Schematic showing, from top
to bottom, the targeting construct for the Sox10 
K2 mutation, the
Sox10 wild-type locus and the mutant locus before and after Cre
recombination. The Sox10 exons (I-V) and the mutant Sox10
K2 open reading frame (ORF) are shown as boxes, the 4.5 kb and
1.5 kb flanking regions as bars. Regions of homology between wild-type locus
and targeting vector are depicted as thick black lines, introns III and IV as
thin open boxes and surrounding genomic regions not contained in the targeting
construct as dashed lines. Plasmid backbone sequences of the targeting
construct are indicated by a thin line. Restriction sites for NcoI
(N), BamHI (H) and ScaI (S) are shown, as are the locations
of the 5' and 3' probes and the start codon of the Sox10
gene (ATG). The arrowheads indicate the locations of primers 1,2,3,4 used for
quantitative RT-PCR. neo, neomycin resistance cassette; loxP, recognition
sites for Cre recombinase; Tk, herpes simplex virus thymidine kinase gene
cassette. (B) Schematic of the mutant locus for the Sox10 aa1 mutation.
Recombination into the Sox10 genomic locus was as depicted for the
Sox10 K2 construct. (C) Southern blot analysis of DNA from
wild-type (wt) and heterozygous (+/aa1 and +/K2) ES cells digested with
NcoI for use with the 5' probe, and with
BamHI/ScaI for the 3' probe. The size of bands
corresponding to the wild-type (6.6 kb for the 5' probe and 4.6 kb for
the 3' probe) and the targeted alleles (10.9 and 5.9 kb, respectively,
for the 5' probe; 10.2 and 10.4 kb, respectively, for the 3'
probe) are indicated. (D) PCR genotyping of wild-type (wt),
heterozygous (+/K2 and +/aa1) and homozygous (K2/K2 and
aa1/aa1) mouse embryos at 18.5 dpc. DNA fragments in the size marker (M) are
1.0 kb and 0.5 kb.
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