The targeting construct contains a 3.1 kb En2 5' fragment, an Frt-flanked PGK-neo cassette followed by a LoxP site in the En2 intron and a 4.8 kb En2 genomic fragment containing a LoxP site in the XbaI site of the 3' UTR. Flpe-mediated recombination deleted the PGK-neo, producing an En2 allele with a floxed region. (J:156169)