A loxP flanked targeting cassette containing neo, URA and tACE-Cre recombinase, replaced 60 bp of a conserved praline-rich domain in the Gnasxl-specific exon. A deletion of only 60 bp was unlikely to disrupt imprinting at the complex Gnas locus. Southern blot confirmed recombination. RT-PCR indicated lack of mutant transcript after paternal transmission as Cre expression excised the cassette in the male germ line. Gnas and exon 1A transcripts were not affected in mutants as shown by RT-PCR. Northern blot demonstrated that Gnas transcript levels are also normal in mutants.