|
| Status |
Public on Dec 11, 2014 |
| Title |
8 weeks control, replicate 4 |
| Sample type |
SRA |
| |
|
| Source name |
Liver, control, 8 weeks
|
| Organism |
Mus musculus |
| Characteristics |
strain/background: C57BL/6
tissue: liver (bottom half of right lobe)
duration of huri expression: 8 weeks
ectopic expression: none
|
| Treatment protocol |
No treatments were done, except hURI ectopic expression since conception.
|
| Growth protocol |
Bottom half of the right liver lobe was taken for each experiment.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Freshly taken liver tissues were snap frozen in liquid nitrogen, lysed in 1.5 ml of TRIZOL solution using precellys homogenizer at 25,000 rpm for 30 s. The mixture was mixed vigorously by vortexing. 0.4 volume of chloroform was added, homogenised by hand for 15 s and incubated at room temperature for 5 min. Then, a centrifugation step for 15 min at 12,000 speed was performed and the upper aqueous phase was transferred into a fresh Eppendorf tube containing 60% volume of Trizol. The upper phase was collected and a second chloroform extraction step was performed. Subsequently, RNA was precipitated by adding 0.8 volumes of 100% isopropanol to the aqueous phase, mixed and incubated at room temperature for 10 min. The RNA-isopropanol mixture was centrifugated for 13.000 rpm for 20 min and total RNA was precipitated. The supernatant was decanted and the pellet was washed with freshly prepared 80% ethanol.
1.5 µg of total RNA samples was used as provided by the user. RNA Integrity Numbers were 8.3 on average (range 7.5 to 9.2) when assayed by Lab-chip technology on an Agilent 2100 Bioanalyzer. PolyA+ RNA fraction was extracted and randomly fragmented, converted to double stranded cDNA and processed through subsequent enzymatic treatments of end-repair, dA-tailing, and ligation to adapters as in Illumina's "TruSeq RNA Sample Preparation Guide" (Part # 15008136 Rev. A). Adapter-ligated library was completed by limited-cycle PCR with Illumina PE primers (10 cycles). The resulting purified cDNA library was applied to an Illumina flow cell for cluster generation (TruSeq cluster generation kit v5) and sequenced on the Genome Analyzer IIx with SBS TruSeq v5 reagents by following manufacturer's protocols.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Description |
Isolation of polyA+ RNA from total RNA.
|
| Data processing |
Read files were quality-checked with FastQC.
Reads were aligned with the mouse genome (NCBI37/mm9) with TopHat-1.3.1 (using Bowtie 0.12.7 and Samtools 0.1.16), allowing 2 mismatches and 5 multihits.
Transcripts quantification and differential expression calculated with Cufflinks 1.3.0.
Transcripts annotations used were Mus musculus NCBIM37.65 from Ensembl.
Genome_build: MGSCv37
Supplementary_files_format_and_content: Tables were generated with Cufflinks 1.3.0 and contain transcripts abundance measured in FPKM values.
|
| |
|
| Submission date |
Jul 09, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Nabil Djouder |
| E-mail(s) |
ndjouder@cnio.es
|
| Organization name |
CNIO
|
| Lab |
Growth Factors, Nutrients and Cancer
|
| Street address |
C/Melchor Fernandez Almagro, 3
|
| City |
Madrid |
| ZIP/Postal code |
28029 |
| Country |
Spain |
| |
|
| Platform ID |
GPL11002 |
| Series (1) |
| GSE48654 |
Genes regulated in mouse liver by expressing ectopically hURI in hepatocytes |
|
| Relations |
| BioSample |
SAMN02230251 |
| SRA |
SRX319219 |
