A cre/EGFP fusion gene and Frt-flanked hygro selection cassette were inserted using recombineering into an Nkx2-2 basal exchange vector such that cre/EGFP was fused in-frame with the ATG start codon, replacing all but 25 base pairs of the first coding exon. This vector was co-injected with a cre-expression vector into ES cells containing an Nkx2-2 (loxed cassette acceptor) allele. Correctly exchanged ES clones were used to produce chimeras which were bred to B6 mice to verify germline transmission. Animals were bred to ACTB-Flpe mice to excise the hygromycin marker.