Figure 1.

OGT mutagenesis in ES cells and mice. A mouse OGT genomic clone bearing five exons (filled boxes) was used to construct two types of gene-targeting vectors. (A) The first vector would be expected to produce a null mutation by deletion of two OGT exons with insertion of a neomycin phosphotransferase cistron (neo). (B) The second vector incorporates loxP sites that initially would generate the OGT^F[tkneo] allele. (C) ES cells bearing the OGT^F[tkneo] allele that are subjected to transient Cre expression should yield both OGT^Δ (type I) and OGT^F (type II) alleles. (D) Homologous recombinants were obtained with the loxP-containing vector shown in B and were characterized by Southern blotting with probes of OGT genomic (Left) or loxP (Right) sequences. (E) Only OGT^F alleles are present among subclones, as represented, in over 200 analyzed specimens. (F) The OGT^Δ allele is detectable by PCR in polyclonal ES cell extracts only at early times after Cre expression. (G) OGT alleles in offspring of chimeric male (♂) mice bred to C57BL/6 females (♀) were analyzed by Southern blotting. Female offspring are either homozygous wild type or heterozygous for the OGT^F allele, whereas male offspring are either homozygous wild type or appear homozygous for the OGT^F allele. wt, wild type.