|
| Status |
Public on Jun 30, 2014 |
| Title |
aorta_M_placebo_rep4 |
| Sample type |
RNA |
| |
|
| Source name |
aorta, placebo
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
genotype/variation: ApoE deficient
gender: female (ovariectomized)
age: 4-5 weeks
agent: placebo
tissue: aorta
diet: high-fat Western-type diet
|
| Treatment protocol |
Female ApoE-deficient mice were ovariectomized at the age of 4 - 5 weeks and two weeks later implanted subcutaneously with pellets (Innovative Research of America) constantly releasing the respective hormone (medroxyprogesterone acetate, norethisterone acetate or placebo) for 90 days. For the time of hormone substitution mice were fed a high fat diet. After 78 - 90 days of treatment mice were sacrificed and aortas removed from behind the aortic arch curvature to approximately 1 mm before the vessel-branches to the kidney and snap-frozen in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Tissue samples were pulverised, immediately transferred into 1 ml of peqGOLD TriFastTM (peqlab), incubated at room temperature for 1 hour and vortexed every 10 minutes. Extraction of RNA was then performed according to the manufacturer`s instructions. Final purification of total RNA was performed according to the RNET-Asy Mini Kit RNA cleanup protocol (Qiagen)
|
| Label |
Cy3
|
| Label protocol |
Synthesis of cDNA and subsequent fluorescent labeling of cRNA was performed according to the manufacturers´ protocol (One-Color Microarray-Based Gene Expression Analysis / Low Input Quick Amp Labeling; Agilent Technologies). Briefly, 100 ng of total RNA were converted to cDNA, followed by in vitro transcription and incorporation of Cy3-CTP into nascent cRNA
|
| |
|
| Hybridization protocol |
After fragmentation labeled cRNA was hybridized to Agilent 4x44k Whole Mouse Genome v1 Microarrays for 17 h at 65 °C according to the manufacturers´ protocol (One-Color Microarray-Based Gene Expression Analysis, Low Input Quick Amp Labeling, Agilent Technologies)
|
| Scan protocol |
Microarray slides were scanned as described in the manufacturers´ protocol (Scanner G2505C, Agilent Technologies).
|
| Description |
gene expresion after 90 days of treament
AP8_US45103065_251486819714_S01_GE1_107_Sep09_1_4
|
| Data processing |
Signal intensities on 20 bit tiff images were calculated by Feature Extraction software (FE, Vers. 10.7.1.1 / 11.0.1.1; Agilent Technologies) using scan protocol GE1_107_Sep09 / grid 014868_D_F_20100123 (medroxyprogesterone acetate experiments) and scan protocol GE1_1100_Jul11 / grid 014868_D_F_20130207 (norethisterone acetate experiments), respectively. Data analyses were conducted with GeneSpring GX software (Vers. 12.5; Agilent Technologies). Probe signal intensities were quantile normalized across all samples to reduce inter-array variability. Input data pre-processing was concluded by baseline transformation to the median of all samples
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| |
|
| Submission date |
May 27, 2014 |
| Last update date |
Jun 30, 2014 |
| Contact name |
Karl Koehrer |
| E-mail(s) |
rene.deenen@hhu.de
|
| Organization name |
University of Duesseldorf
|
| Department |
BMFZ
|
| Lab |
GTL
|
| Street address |
Universitaetsstr. 1
|
| City |
Duesseldorf |
| ZIP/Postal code |
40225 |
| Country |
Germany |
| |
|
| Platform ID |
GPL7202 |
| Series (1) |
| GSE57972 |
Analysis of aortic gene expression in ovariectomized ApoE-deficient mice substituted with medroxyprogesterone acetate, norethisterone acetate or placebo |
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