|
| Status |
Public on Oct 02, 2011 |
| Title |
colon_heme_repl6 |
| Sample type |
RNA |
| |
|
| Source name |
colon, 2 weeks of heme diet (40en%fat and 0.5 umol/g heme added)
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
gender: male
age: 11 weeks
tissue: colon
treatment: high fat and low calcium diet with heme
treatment duration: 14 days
|
| Treatment protocol |
Male control and heme-fed mice were maintained on a high fat-low calcium diet (high fat and low calcium diet with or high fat and low calcium diet without heme) for 2 weeks. After 2 weeks diet intervention, all mice were sacrificed.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Colons were taken out of the mice and mesenteric fat was removed. Colons were cut open longitudinally, washed in PBS and scraped. RNA was isolated from scrapings. All mice were individually analyzed by microarray.
Total RNA was prepared from mouse colon scrapings using TRIzol reagent, and total RNA was purified using Qiagen RNeasy columns. RNA integrity was checked on chip analysis (Agilent 2100 bioanalyzer, Agilent Technologies, Amsterdam, the Netherlands) according to the manufacturer's instructions. RNA was judged as suitable for array hybridization only if samples exhibited intact bands corresponding to the 18S and 28S ribosomal RNA subunits, and displayed no chromosomal peaks or RNA degradation products (RNA Integrity Number > 8.0).
|
| Label |
biotin
|
| Label protocol |
The Affymetrix GeneChip RNA One cycle Amplification Kit was used to prepare labelled cRNA from 5 μg of total RNA. The protocol was conducted using the reagents provided by Affymetrix in the One-Cycle cDNA synthesis kit (P/N 900431), One-Cycle IVT labelling kit (P/N 90449) and GeneChip Sample Cleanup Module (P/N 900371). A detailed description can be found in the Genechip Expression Analysis Technical Manual, section 2 (Eukaryotic Sample and Array Processing), chapter 1 (Eukaryotic Target Preparation) (P/N 701025, revision 6).
|
| |
|
| Hybridization protocol |
Hybridization of 10ug cRNA was done overnight for 16 hours at 45ºC in a Hybridization Oven 640 (Affymetrix). The protocol is conducted as described in the Genechip Expression Analysis Technical Manual, section 2 (Eukaryotic Sample and Array Processing), chapter 2 (Eukaryotic Target Hybridization) (P/N 701027, revision 5).
|
| Scan protocol |
Arrays were scanned on an Affymetrix 3000 7G scanner, as described n the Genechip Expression Analysis Technical Manual, section 2 (Eukaryotic Sample and Array Processing), chapter 2 (Eukaryotic Arrays: Washing, Staining and Scanning (P/N 701028, revision 5).
|
| Description |
A108_C12_HEME_MCOLON
Colon from heme-fed mouse after 2 weeks of heme diet (40en%fat and 0.5 umol/g heme added).
|
| Data processing |
Expression estimates were calculated using GCRMA (v2.22.0) in Bioconductor, applying the empirical Bayes (EB) model for background estimation.
|
| |
|
| Submission date |
Mar 08, 2011 |
| Last update date |
Oct 03, 2011 |
| Contact name |
Guido Hooiveld |
| E-mail(s) |
[email protected]
|
| Organization name |
Wageningen University
|
| Department |
Div. Human Nutrition & Health
|
| Lab |
Nutrition, Metabolism & Genomics Group
|
| Street address |
HELIX, Stippeneng 4
|
| City |
Wageningen |
| ZIP/Postal code |
NL-6708WE |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL1261 |
| Series (2) |
| GSE27848 |
Dietary heme stimulates epithelial cell turnover by downregulating feedback inhibitors of proliferation in murine colon (part 2) |
| GSE27849 |
Dietary heme stimulates epithelial cell turnover by downregulating feedback inhibitors of proliferation in murine colon |
|
