|
| Status |
Public on Mar 15, 2019 |
| Title |
Couple 3 Female Cftr N/N ileum |
| Sample type |
SRA |
| |
|
| Source name |
Small intestine, <6 cm proximal to the ileocecal valve
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Ileum - Whole tissue section
gender: F
strain: FVB
genotype: Cftr N/N
|
| Treatment protocol |
Laxative treatment was discontinued >4 days before sampling. Where indicated (ABX) the drinking water was supplemented with ciprofloxacin (0.3 g/L) and metronidazole (0.5 g/L).
|
| Growth protocol |
Animals were reared on a low fiber diet (C1013; Altromin) and an osmotic laxative (ad libitum)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
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).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Ion Torrent Proton |
| |
|
| Description |
Littermate couple 3_f Cftr WT ileum
|
| Data processing |
Basecalls performed using Illumina's Real Time Analysis Software (HiSeq2000 ) or Ion Proton use Proton™ Torrent Server and Torrent Suite Software
Raw reads are filtered by removing the adaptors, reads in which unknown bases are more than 10%, low quality reads (the percentage of low quality bases is over 50% in a read, where the low quality base is defined as base whose sequencing quality is no more than 5). After filtering, the remaining reads are stored as FASTQ.
Reads are mapped and RPKM values are calculated by CLC Genomics workbench 7.5.1, using default settings
Genome_build: GRCm38.76
Supplementary_files_format_and_content: Tab-delimited text file with normalized RPKM values for all the samples
|
| |
|
| Submission date |
Dec 29, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Marcel Bijvelds |
| Organization name |
Erasmus MC University Medical Center
|
| Department |
Gastroenterology and Hepatology
|
| Street address |
PO Box 2040
|
| City |
Rotterdam |
| ZIP/Postal code |
3000CA |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL18635 |
| Series (1) |
| GSE92991 |
Gene expression in the ileum of Cftr null mice |
|
| Relations |
| BioSample |
SAMN06189446 |
| SRA |
SRX2449440 |
