GSM2441724: Couple 3 Female Cftr N/N ileum; Mus musculus; RNA-Seq
Sampling from a CF animal and a sex-matched littermate control was performed within a time window of 20 min, and between 12:00-14:00h. Tissue was homogenized with a rotor-stator homogenizer in Trizol reagent (Qiagen), and total RNA was extracted using the Nucleospin RNA kit (Macherey-Nagel). Integrity of the extracted RNA was verified by gel electrophoresis. Libraries were prepared by BGI. In brief, the mRNA is enriched by using the oligo(dT) magnetic beads. Mixed with the fragmentation buffer, the mRNA is fragmented into short fragments. Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3’-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are enriched by PCR amplification. During the QC step, Agilent 2100 Bioanaylzer and ABI StepOnePlus Real-Time PCR System are used to qualify and quantify of the sample library. The library products are ready for sequencing via Illumina HiSeqTM 2000 or Ion Proton System (4476610).
GEO Accession
GSM2441724