Rosa lsl dtr

The Lgr5-EGFP-IRES-CreERT2, Hopx-CreERT2, Hopx-GFP, Rosa-LSL-DTR and Rosa-LSL-tdTomato mice were purchased from Jackson Laboratory. The Axin2-CreERT2-tdTomato line was a kind gift from Dr. Edward E. Morrisey (University of Pennsylvania). Male mice 8-12 weeks of age were used to perform experiments. Detailed information on generation and genotyping of those mouse lines has been published previously (Barker et al., 2007 (link), Buch et al., 2005 (link), Frank et al., 2016 (link), Takeda et al., 2011 (link), Takeda et al., 2013 (link)). All animal work was performed in accordance to the protocols approved by the Washington University School of Medicine Animal Studies Committee.

All mice were maintained on the C57BL/6 background. C57BL/6 mice, OTII T-cell receptor transgenic mice (25 (link)), Foxp3^GFPCreERT2 mice (26 (link)), Rosa^lslDTR (27 (link)), were purchased from The Jackson Laboratory (Bar Harbor, ME). All mice were fed a routine chow diet. Co-housed littermates were used for experimental controls. All mice were weaned at DOL 21. Adult mice were 8 to 16 weeks of age when analyzed unless stated otherwise. Foxp3^GFPCreERT2 mice and Rosa^lslDTR mice were bred for Foxp3^GFPiDTR mice, which were injected with 100µg tamoxifen (Sigma-Aldrich, St. Louis, MO, USA) dissolved in sunflower seed oil with 20% ethanol (Sigma-Aldrich) intraperitoneally (i.p.) on DOL 14 or 24. Mice were then aged to 8 weeks old (DOL 56), and injected with 50µg/kg diphtheria toxin (DT) (Sigma Aldrich) i.p. In some experiments to validate deletion mechanism, Foxp3^GFPiDTR mice were injected with tamoxifen on DOL21, and diphtheria toxin (DT) on DOL24. For controls, C57Bl/6 mice were injected with tamoxifen or vehicle (sunflower seed oil with 20% ethanol) on DOL24, and DT on DOL56 to minimize variation due to treatments. Animal procedures and protocols were performed in accordance with the Institutional Animal Care and Use Committee at Washington University School of Medicine.