GSM8833212: Duo WT rep 7; Mus musculus; RNA-Seq
The small intestine was extracted in its entirety from stomach to cecum and flushed with cold phosphate buffered saline (PBS). Three-inch long intestinal segments were collected from the beginning, middle and end of the small intestine to correspond with duodenum, jejunum, and ileum respectively and frozen immediately in liquid nitrogen. The jejunum section was divided in half lengthwise prior to freezing, using one section for whole tissue RNA and the other for crypt isolation and subsequent RNA extraction. RNA was isolated using Rneasy mini kit (Qiagen), concentration and quality checked with the Nanodrop (ThermoScientific). RNA-seq libraries were generated using TruSeq stranded Total RNA Ribo-Zero mouse gold kit (Illumina, San Diego, CA) and the quality of resulting libraries was assessed using Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA). Ileum (WT n=4, CF n=5) sequencing was carried out using an Illumina HiSeq 2500 Rapid Run flow cell – 2x100bp run. Duodenum (WT n=9, CF n=9), jejunum (WT n=8, CF n=9) and jejunum crypts (WT n=8, CF n=9) sequencing was carried out using an Illumina NextSeq 550 HO flowcell – 2x75bp run