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Fig. 1. Generation of the Myogflox allele, targeted ES cells
and heterozygous mice. (A) The top line shows the myogenin locus
with its three exons (black rectangles). The second line shows the targeting
construct. The region of homologous DNA (thicker line) spans 3.8 kb and is
flanked by EcoRI sites. Also shown are loxP sites (red
triangles) and a neomycin cassette placed into the BamHI site in the
first intron (gray rectangle). A thymidine kinase (TK) cassette (white
rectangle) lies upstream of the homology region. The next two lines depict the
targeted Myog locus before and after Cre recombinase-mediated
deletion of the first exon and neomycin cassette. (B) Southern genome
hybridization of ES cell DNA digested with BamHI and
HindIII, confirming proper targeting. A 3' probe derived from
the third exon immediately 3' to the EcoRI site outside of the
homology region was used in the hybridization analysis. The 2 kb band shown on
the figure represents the wild-type BamHI fragment bounding the
second and third exons (Myog+). The 6 kb band represents
the targeted BamHI-HindIII fragment
(Myogflox) containing the neomycin cassette. The left and
right lanes show DNA from targeted and wild-type ES cell lines, respectively.
(C) Southern genome hybridization of tail DNA digested with
EcoRI and hybridized with a Myog cDNA probe. Tail DNA was
obtained from P10 pups resulting from mating Myog+/- and
Myog+/flox mice. The left, idle and right lanes show DNA
from Myog+/flox, Myog+/- and
Myogflox/- mice, respectively. The 2 kb band representing
the Myog- allele is the result of a EcoRI site in
the neomycin cassette that is not present in the neomycin cassette of the
Myogflox allele. The origins of the 4 kb and 6 kb bands
associated with the wild-type and Myogflox alleles,
respectively, are shown in A.
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