title: Illumina HiSeq 2500 sequencing; Bulk RNA-seq of Adult and Fetal Enteroids (Control or Treated with DBZ)
library_construction_protocol: Fresh human fetal epithelium was isolated and maintained as previously described (Tsai et al. 2018, Supplemental Protocol 1). Once enteroids were established, healthy cystic enteroids were manually selected under a stereoscope, bulk-passaged through a 30G needle, and embedded in Matrigel(Corning, 354234). Control culture media consisted of 50% LWRN conditioned media (Miyoshi & Stappenbeck, 2013) and 50% Human 2X basal media [Advanced DMEM/F12(Gibco, 12634-028); Glutamax 4 mM (Gibco, 35050-061); HEPES 20 mM (Gibco, 15630-080); N2 Supplement (2X) (Gibco, 17502-048), B27 Supplement (2X) (17504-044), Penicillin-Streptomycin (2X) (Gibco, 15140-122), N-acetylcysteine(2 mM) (Sigma, A9165-25G), Nicotinamide (20 mM) (Sigma, N0636-061)]. NOTCH inhibition media was control media with the addition of 5 μM DBZ Dibenzazepine; Apexbio Technology LLC; YO-01027). "Enteroids were grown in either control media as mentioned in \""growth protocol\"" and experimental group was treated with 5 μM DBZ in control media. " RNA from each sample was isolated using MagMAX-96 Total RNA (AM1830; Applied Biosystems), RNA isolation kits.  Input for library generation with Takara SMARTer Stranded Total RNA Sample Prep Kit (634876; Takara Bio USA)