Figure 1.

Targeted duplication of the murine Npr1 gene. (A) Homologous recombination and targeting construct. The endogenous Npr1 locus is shown schematically in the top line. The duplication targeting construct (middle line) uses a 6.5-kb 5′ region of homology from immediately upstream of the Npr1 coding region. The 3′ region of homology is a 1.3-kb fragment located approximately 6 kb downstream of exon 22. A 24-kb gap (indicated) is repaired during the recombination and a single crossover event leads to a 32-kb duplication, which includes the complete Npr1 gene and upstream and downstream flanking sequences. The bottom line shows the resultant locus with the two copies of Npr1 in tandem arrangement separated by the neomycin-resistant gene (NEO) (not drawn to scale). ×, XbaI sites; ∗, position of the probe. (B) Southern blots. Correctly targeted clones are identified after digesting ES cell DNA with XbaI and hybridizing with a probe corresponding to exon 1 of Npr1. The probe hybridizes to both 10.4- and 25-kb bands in the duplicated chromosome, but only to a 10.4-kb band in the unmodified chromosome.