A loxP site was inserted into a Ssp I restriction site 1.1 kb upstream of exon 1, and a floxed neomycin cassette was placed 1 kb downstream of exon 1. ES cells with the targeted locus were transiently infected with a cre-expressing adenovirus to generate exon 1 flanked with loxP sites. Expression of the locus was not affected as determined by immunoblot analysis of spleen, thymus, brain and lung tissues.