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Status |
Public on May 12, 2010 |
Title |
IRA_12 |
Sample type |
RNA |
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Source name |
Infrarenal region of abdominal aorta
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Organism |
Mus musculus |
Characteristics |
ApoE-/- (C57B6 background), male mouse, 14 weeks old
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Biomaterial provider |
Animal Resource Centre (http://www.arc.wa.gov.au/) PO Box 1180, Canning Vale WA 6970, Australia
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Extracted molecule |
total RNA |
Extraction protocol |
RNA was extracted from excised artery segments by tissue disruption in Tri-Reagent (Sigma) and extraction as per manufacturers instructions. Isolated RNA was further purified on RNeasy columns (Qiagen) as per manufacturers instructions. The concentration and quality of total RNA was measured by spectrophotometry at OD260/280 and the quality of the total RNA sample was assessed using an Agilent Bioanalyser and Agilent RNA6000 Nano Lab Chip.
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Label |
biotin, Cy5-Streptavidin
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Label protocol |
Biotin-labeled cRNA was prepared by linear amplification of the Poly(A)+ RNA population within the total RNA sample. Briefly, 0.51 µg of total RNA was reverse transcribed after priming with a DNA oligonucleotide containing T7 RNA polymerase promoter 5' to a d(T)24 sequence. After second-strand cDNA synthesis and purification of double-stranded cDNA, in vitro transcription was performed using T7 RNA polymerase in the presence of biotinylated UTP.
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Hybridization protocol |
10 µg of purified cRNA was fragmented to uniform size and applied to CodeLink Bioarrays (GE Healthcare) in hybridization buffer. Arrays were hybridized at 37ºC for 18 hrs in a temperature-controlled shaking incubator. Arrays were washed in 0.75x TNT at 46º C for 1 hr and stained with Cy5-Streptavidin dye conjugate for 30 min.
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Scan protocol |
Arrays were scanned with a GenePix™ 4000B scanner.
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Description |
-
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Data processing |
Array data was processed with CodeLink Expression Analysis software (GE Healthcare) and data was analyzed with GeneSpring software (Silicon Genetics). To compare individual expression values across arrays, raw intensity data (generated from CodeLink Expression 2 software) from each gene was normalized to the median intensity of the array. Only genes which had values greater than the background intensity in at least one condition were used for further analysis.
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Submission date |
Jan 29, 2007 |
Last update date |
May 04, 2009 |
Contact name |
Jonathan Golledge |
E-mail(s) |
jonathan.golledge@jcu.edu.au
|
Phone |
+67 7 47814730
|
Fax |
+61 7 4781 6986
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URL |
http://www.jcu.edu.au/school/medicine/VBU.htm
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Organization name |
James Cook University
|
Department |
School of Medicine
|
Lab |
Vascular Biology Unit
|
Street address |
School of Medicine
|
City |
Townsville |
State/province |
Queensland |
ZIP/Postal code |
4811 |
Country |
Australia |
|
|
Platform ID |
GPL2897 |
Series (1) |
GSE7006 |
Comparison of infrarenal and suprarenal sections of mouse ApoE-/- abdominal aorta |
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