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Fig. 1. Targeting of the LRE and RARE in the mouse Cdx1 promoter.
(A) Schematic of the wild-type (WT) Cdx1 locus, targeting
vector, targeted allele and Cre-recombined allele. Probe A (5'
external probe) was used to screen for targeted ES cells, probe B (5'
internal probe) was used to confirm the predicted targeting event and probe C
(3' internal probe) was used to confirm excision of the floxed neomycin
selection cassette. E, EcoRI; H, HindIII; K, KpnI;
S, SacI; E1, Exon 1; *, mutated RARE; ++, mutated LRE.
(B) Southern blot analysis of DNA from wild-type (left), heterozygous
targeted (middle) and heterozygous Cre-recombined (right) offspring
using probe B and the indicated restriction endonucleases (see A). Concomitant
integration of mutated LRE and LRE+RARE within the targeted allele was
determined by the introduction of novel restriction sites: for the RARE
mutation, a SacI restriction site was observed by Southern blot
analysis (last lane of middle and right panels); for the LRE mutation, novel
SfuI restriction sites were assessed by restriction of a PCR product
spanning the LRE (lower left panel; see also Materials and methods).
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