Briefly, the targeting vector (shown in Figure 2A) places a loxP site within intron 18 and a loxP-flanked MC1neopA selection cassette in intron 17 of the Atrx gene. A detailed description of the targeting construct is provided in [20]. Linearised plasmid (150 μg) was electroporated into 1 × 108 E14Tg2a ES cells, and colonies resistant to G418 and ganciclovir were isolated. Homologous targeting events were identified by Southern blot of EcoRI-digested DNA and hybridisation with a 5′ probe (generated with primers PPS1.20 and PPS1.27) and a 3′ probe (a 0.9-kb HaeIII fragment) as shown in Figure 2A and 2B. DNA from correctly targeted clones was also digested with SacI and analysed by Southern blot and hybridisation with a probe from within intron 17 (a PCR product generated with primers PPS1.15 and Xnp46) to confirm that the loxP site within intron 18 had been included within the crossed-over region (Figure 2A and 2C). Sequences of primers are shown in Table S1.