GSM5014477: Intestine 5; Mus musculus; RNA-Seq
Intestines were isolated from mice then flushed with PBS, cut open to lay flat, seperated into duodenum, jejunum, and ileum, and the bottom two cms were scraped to collect the epithelial cells which were then snap frozen in liquid N2. For liver tissue, the liver was perfused, then 50 mg of tissue was cut and snap frozen in liquid N2 Sequenced RNA samples first passed quality control using the RNA tape on Tapestation 4200 (Agilent) and were quantified with Qubit Fluorometer (Thermo Fisher). Only those samples that passed quality control were used for analysis. The RNA libraries were prepared with QIAseq Stranded Total RNA library Kit (Qiagen) using 100 ng input RNA. Oligo-dT probes attached to Pure mRNA beads were used to enrich for mRNA (Qiagen). RNA was fragmented, reverse transcribed into cDNA, and ligated with Illumina sequencing adaptors. The size selection for libraries was performed using SPRIselect beads (Beckman Coulter) and purity of the libraries was analyzed using the DNA 1000 tape Tapestation 4200 (Agilent). The prepared libraries were pooled and sequenced using NextSeq 500 (Illumina), generating ~20 million 2×76 bp paired-end reads per samples. Two runs were performed on the intestine tissue samples and merged for analysis.