|
| Status |
Public on Dec 24, 2008 |
| Title |
MG_An_50 |
| Sample type |
RNA |
| |
|
| Source name |
Bilateral thoracical C57BL/6 mouse mammary gland
|
| Organism |
Mus musculus |
| Characteristics |
Diet regimen: food consumed ad libitum (AL)
|
| Treatment protocol |
Female C57BL/6 mice (9 weeks old) were randomized into 4 experimental groups: 1) food consumed ad libitum (AL); 2) AL with access to running wheels (AL+EX); 3) 30% CR; and 4) 30% CR with access to running wheels (CR+EX). CR mice received 70% of calories but 100% of all other nutrients compared to AL mice. Food consumption, weight gain, and physical activity levels were recorded for 6 weeks. RNA samples from animal mammary glands were analysed by Affymetrix oligo-microarray platform, in particular 10 samples from AL group (control), 9 samples from AL+EX group, 9 samples from CR group and 9 samples from CR+EX group were analysed.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total mammary gland RNA was isolated from pooled left and right thoracic mammary glands (approximately 60 mg each) prior to microarray analysis, using TRIzol extraction reagent (Invitrogen, Rockville, MD) and the RNeasy Midi Cleanup Kit (Qiagen, Valencia, CA) in accordance with the manufacturers’ protocols. Total RNA concentration was determined by spectrophotometric evaluation of absorbance at 260 nm, and RNA integrity was confirmed by 1% agarose gel electrophoresis. RNA quality of random samples was also tested on an Agilent 2100 bio-analyzer (Agilent Technologies, Palo Alto CA).
|
| Label |
biotin
|
| Label protocol |
Affymetrix recommended protocol was followed for labeling (Expression Analysis Technical Manual, 2001, Affymetrix). RNA was biotin-labeled using the Enzo Bioarray-High Yield-RNA Transcript Labeling Kit (Cat #42655-20)
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| |
|
| Hybridization protocol |
Affymetrix recommended protocol was followed for hybridation process. Hybridation cocktail was prepared by mixing Control oligo B2, 20X Euk. Hyb controls, Herring Sperm DNA, Acetylated BSA, 2X Hybridization Buffer and RNase-free Water at the requested concentration. Following fragmentation, 15 ug of cRNA, were hybridized on Affymetrix GeneChip Mouse Expression Array 430A . The microarrays were than incubated for 16 hours (o/n) at 45˚C and 60rpm. (Takahashi et al Using DNA microarray analyses to elucidate the effects of genistein in androgen-responsive prostate cancer cells: identification of novel targets. Mol Carcinog 2004;41:108-19)
|
| Scan protocol |
Affymetrix equipment and recommended protocol were used for the scan process. (Takahashi et al Using DNA microarray analyses to elucidate the effects of genistein in androgen-responsive prostate cancer cells: identification of novel targets. Mol Carcinog 2004;41:108-19)
|
| Description |
Effects of calorie restriction and exercise alone or in association in mammary gland gene expression
|
| Data processing |
The signal values and detection calls of the probe sets were determined using Affymetrix GCOS (ver. 1) software. Relative intensity variation across arrays was normalized by scaling to an average signal level of 500 counts, excluding low 2% and high 2% signals. These signal values were transformed to a logarithmic scale (base 2) for statistical calculations.
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| |
|
| Submission date |
Dec 23, 2008 |
| Last update date |
Dec 23, 2008 |
| Contact name |
Jackie Lavigne |
| Organization name |
NCI/NIH
|
| Department |
DCEG
|
| Street address |
6120 Executive Blvd. Rm. 6114
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL339 |
| Series (1) |
| GSE14202 |
Effects of calorie restriction and exercise on mammary gland gene expression in C57BL/6 mice |
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