Lgr5 egfp ires creert2 mice

C57BL/6J wild type (WT) mice and B6129P2-Lgr5^{tm1(cre/ERT)Cle}/J mice with Lgr5-EGFP expression confined to ISC (hereafter referred to as Lgr5 TG mice) were purchased from the Jackson Laboratory (Bar Harbor, ME). To generate B6129P2-Lgr5^{tm1(cre/ERT)Cle}/J;Villi-HB-EGF/+ mice (Lgr5 TG mice that overexpress HB-EGF in the intestines; hereafter referred to as Lgr5-HB-EGF TG mice), B6129P2-Lgr5^{tm1(cre/ERT)Cle}/J mice were crossed with homogeneous transgenic Villi-HB-EGF/Villi-HB-EGF mice that we produced in our laboratory.²⁷ In addition, Lgr5-lacZ mice were generated by crossing Lgr5-EGFP-IREScreERT2 mice to Rosa26-lacZ Cre-reporter mice, both obtained from Jackson Laboratory (Bar Harbor, ME).^{20 (link)} One to two month old mice were injected intraperitoneally (IP) with tamoxifen (200 μl;10 mg/ml) in sunflower oil to induce LacZ expression in Lgr5⁺ ISC and their progeny cells.

All mice including Lgr5-EGFP-ires-creERT2 mice, TCF/LEF-reporter mice (B6.Cg-Tg (BAT-lacZ)3Picc/J), and C57BL/6j mice at 6–12 weeks of age were obtained from Jackson Laboratories. All animal procedures were approved by the University of Louisville Institutional Animal Care and Use Committee. Mice were administered by gavage EPDENs resuspended in PBS as described previously[25 (link)].

All studies involving mice were approved by the Stony Brook University Institutional Animal Care and Use Committee (IACUC) and maintained on a 12:12 hr light-dark cycle. C57BL/6J mice carrying Klf5 alleles flanked by loxP sites (Klf5^fl/fl) were previously described [29 (link)] and graciously provided by Dr. Ryozo Nagai. C57BL/6 mice carrying the inducible Cre recombinase gene under regulation of the Lgr5 promoter (Lgr5-EGFP-IRES-CreER^T2 mice; henceforth designated as Lgr5-Cre) were purchased from Jackson labs (ME, USA). To establish Lgr5-EGFP-IRES-CreER^T2/Klf5^fl/fl (designated as Lgr5-Klf5^fl/fl) mice, Klf5^fl/fl mice were initially crossed with Lgr5-Cre mice and its progeny then backcrossed to Klf5^fl/fl mice. For all experiments, Lgr5-Cre mice were used as controls. tamoxifen treatment of mice was per protocol previously described [30 (link)] (Suppl. Fig. 1). Eight week old Lgr5-Klf5^fl/fl and Lgr5-Cre mice were injected intraperitoneally (I.P.) with 1 mg of tamoxifen (10 mg/ml, dissolved in sterile corn oil, Sigma, MO, USA) for five consecutive days. Animals were sacrificed on days 3, 5, 7 and 11 after the start of tamoxifen injections for short-term (acute) tamoxifen-treatment study (Suppl. Fig. 1A) and on days 14, 28, 56 and 112 for the long-term (chronic) tamoxifen-treatment study (Suppl. Fig. 1B).