A loxP site was inserted upstream of exon 1 (GRCm38; Chr7:127841278) and a second loxP site, followed by the phosphoglucokinase promoter-driven neomycin resistance gene (PGK-Neo) flanked by two FRT sites, was introduced in intron 4 (GRCm38; Chr7:127842997). The neomycin selection cassette was removed via Flp-mediated recombination, leaving exons 1-4 floxed. Subsequent Cre expression in the germline resulted in the deletion of the loxP-flanked genomic sequences, generating a knockout allele.