2.6. Mouse Intestinal Crypt Isolation and Organoid Culture

Intestinal crypts from 6–8-week-old Apc^Min/+ mice and wild-type mice (Jackson Laboratory, Bar Harbor, ME, USA) were isolated from tumor-free small intestinal tissues based on a previous study [29]. Crypts were cultured under IntestiCult™ organoid media supplemented with EGF, Noggin, and R-Spondin (Stemcell), and Primocin mixed 1:1 with Growth-Factor-Reduced Matrigel (Corning). An amount of 50 μL of the mixture was plated in a prewarmed 48-well plate and incubated for 10 min at 37 °C. For Gln deprivation experiments, Gln-free DMEM/F-12 (Gibco) was supplemented with L-Glutamine (Corning) or Dimethyl alpha-ketoglutarate (Tokyo Chemical Industry, Tokyo, Japan) as indicated.