U.S. flag

An official website of the United States government

Format

Send to:

Choose Destination

ERX1403328: Illumina HiSeq 2000 paired end sequencing; RNA-Seq analysis of gene expression divergence in mammalian liver
1 ILLUMINA (Illumina HiSeq 2000) run: 29.2M spots, 2.9G bases, 1.7Gb downloads

Design: RNA-Seq analysis of gene expression divergence in mammalian liver
Submitted by: Cambridge Institute - Cancer Research UK and University of Cambridge (Cambridge Institute - Cancer Research UK and Unive)
Study: RNA-Seq analysis of gene expression divergence in mammalian liver
show Abstracthide Abstract
The connection between evolution of regulatory elements and downstream gene expression is incompletely understood in mammals. We carried out a large-scale comparison of matched ChIP-seq and RNA-seq experiments across twenty species of mammals, using liver as a model somatic tissue. Herein we report the RNA-seq dataset, comprising twenty five species of mammals. When analysed alongside our previously reported ChIP-seq experiments, these data reveal a number of insights into how mammalian regulatory evolution propagates into differences in gene expression levels. Please note that all mouse RNA-seq libraries and three out of four human libraries have been previously reported in Array Express submission E-MTAB-4052 and are thus omitted here.
Sample: Sample 52
SAMEA3903368 • ERS1090502 • All experiments • All runs
Library:
Name: Sample 52
Instrument: Illumina HiSeq 2000
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: PAIRED
Construction protocol: Tissue samples for RNA extraction were collected in parallel to previously reported ChIP samples {Villar, 2015}. Small pieces of liver tissue (typically 0.5-2 cubic cms) were snap-frozen in dry-ice or liquid nitrogen and stored at -80C. Cetacean liver samples were typically bigger frozen blocks, and for these RNA extraction was conducted on tissue pieces cut from the central section of the frozen piece, to maximise RNA quality. Total RNA was extracted from snap-frozen liver tissue with RNAeasy Mini Kit (Qiagen). 20 mg of tissue were weighed on dry-ice and immediately homogenized in 600 microliters of RLT buffer containing 10 microliters of beta-mercaptoethanol per mililiter of buffer. Tissue samples were homogenized in a Precellys 24 tissue homogenizer, using settings 5500-2x15-015 and Precellys tubes CK28-R (bertin technology). Liver homogenates were processed according to manufacturers' instructions (Qiagen RNAeasy Mini Kit) and total RNA eluted in 50ul RNAse-free water. 10ug total RNA from each sample were treated with 4 units of Turbo DNase (Ambion AM1907), and total RNA samples were run on an Agilent Bioanalyser (RNA nano chip, Agilent) to check RNA integrity (samples were taken forward if RIN values were above 7). Ribosomal RNA was depleted with Ribo-Zero rRNA removal kit (Epicentre RZC110424) as per instructions from the manufacturer, using 5 ug of DNase-treated total RNA. Strand-specific RNA-depleted RNA-Seq libraries were prepared with a modified version of TruSeq RNA Library Preparation kit (Illumina). Fragmentation and first-strand synthesis of rRNA-depleted RNA samples were according to the Illumina protocol. Second-strand cDNA synthesis was done with SuperScript double-stranded cDNA synthesis kit (Life Technologies) at 16C for two hours, using a 10mM dATP, dCTP, dGTP, dUTP nucleotide mix. cDNA was purified with QIAquick PCR purification kit (Qiagen) and end repair, A-tailing and adaptor ligation were performed with Illumina's protocol. Second-strand degradation was done by treatment with one unit of Uracil N-Glycosylase (Life Technologies) at 35C for 15 minutes, prior to PCR enrichment. Libraries were amplified according to Illumina's protocol for 13 PCR cycles, and cleaned-up with Agencourt AMPure beads (Beckman Coulter) with a 1:1 DNA:beads ratio.
Spot descriptor:
forward101  reverse

Experiment attributes:
Experimental Factor: Tupaia belangeri: organism
Runs: 1 run, 29.2M spots, 2.9G bases, 1.7Gb
Run# of Spots# of BasesSizePublished
ERR133170829,241,7482.9G1.7Gb2017-10-13

ID:
4602108

Supplemental Content

Recent activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...