Bulk RNAseq analysis of E14.5 embryonic bladder urothelium following conditional Tbx2/Tbx3 loss

In this study, we investigated the transcriptional changes occurring in the embryonic bladder urothelium following the deletion or reduction of Tbx2 and/or Tbx3 in epithelial progenitors. To ensure the precise isolation of the urothelial lineage, dissociated bladder cells from pooled mutant and control E14.5 embryos were co-labeled with an antibody against the epithelial marker CDH1 and intrinsic GFP fluorescence.

The resulting GFP+CDH1+ dual-positive cells were isolated using fluorescence-activated cell sorting (FACS). Total RNA was extracted, and poly(A)-selected mRNA libraries were analyzed by bulk strand-specific RNA-sequencing (RNA-seq) to identify downstream target genes and molecular pathways regulated by Tbx2/3 during early bladder development.