FIG. 1.
Targeting of the murine α4 locus. (A) Maps of the targeting construct containing three loxP elements (grey bars) and positive (NEO) and negative (DT-A) selection cassettes; the α4 locus (α4+) including the first two exons (black boxes); the locus after homologous recombination (α4flox) and after Cre-mediated excision (α4Δ). Sites for restriction enzymes HindIII (H), KpnI (K), SphI (S), and XbaI are shown; sites introduced after homologous recombination are in bold typeface. The XbaI restriction fragments that are diagnostic for each of the three alleles are shown. (B) Homologous recombination and Cre-mediated excision at the α4 locus. The α4+ allele in ES cells migrates as a 12-kb SphI (lane 1) or 7.1-kb XbaI (lane 3) fragment, and the α4flox allele migrates as a 5.5-kb SphI (lane 2) or 5.7-kb XbaI (lane 4) fragment. BM cells from α4flox/flox mice were transduced with retroviruses encoding Cre and/or GFP, sorted, and cultured in methylcellulose. XbaI-digested DNAs from MSCViresGFP-transduced α4+/+ colonies (lane 5) are compared to those from CFU-C transduced by MSCViresGFP (lane 6) or MSCV-cre-iresGFP (lane 7). The location of the probe used is indicated by the hatched bar in panel A. (C) BM cells from α4+/+, α4flox/+, and α4flox/flox littermates were stained with PE-conjugated anti-α4 integrin antibody (PS/2) and analyzed by FACS (black histograms). An isotype control FACS profile (dotted line) is included for comparison. Typical FACS profiles for each group of animals are shown.