Analyzing the effect of Lef1 deletion in mouse intestinal organoids with single-cell RNA sequencing

We wanted to assess the role of Lef1 in ex vivo organoids using genetic mouse models of intestinal adenomas and scRNA-seq technology. Tumorigenesis was initiated by inducing Apc mutation in Lgr5+ stem cells. Intestinal cells of Lgr5-CreERT;Apc fl/fl (LApc) mouse and Lgr5-CreERT;Apc fl/fl; Lef1 fl/fl (LApcL) mouse were used to generate adenoma organoids. Organoids were cultured without growth factors for three passages and dissociated with Tryple express. We used WT mice as a control to distinguish adenoma cells. WT organoids were cultured with growth factors. Overall design: Single-cell RNA sequencing was performed on mouse intestinal organoids. We deleted Apc with and without Lef1 in the Lgr5+ stem cells of Lgr5-EGFP-IRES-CreERT2;Apcfl/fl;Lef1fl/fl (LApcL) or Lgr5-EGFP-IRES-CreERT2;Apcfl/fl (LApc) mice in vivo and generated organoid cultures. WT mouse organoids were used as a control.