|
| Status |
Public on Apr 05, 2016 |
| Title |
ColonicEpithelium_HighProteinDiet_rep3 |
| Sample type |
RNA |
| |
|
| Source name |
Colonic Epithelium,High Protein Diet,rep3
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: wistar
tissue: colonic epithelium
gender: male
|
| Extracted molecule |
total RNA |
| Extraction protocol |
cRNA synthesis, in vitro transcription, labelling and microarray hybridization were performed as described by manufacture’s protocol (Agilent Technologies). In brief, total RNA was first purified using QIAGEN RNeasy Mini Kit (QIAGEN). For each sample, 2 μg RNA was used for reverse transcribed with T7 Promotor primer, second strand synthesis, and cRNA synthesis, and cRNA purification, to produce high quality cRNA.
|
| Label |
Cy3
|
| Label protocol |
4 μg cRNA were labelled with fluorescent dye Cy3 and purified. 875 ng of each Cy3 labeled cRNA in a volume of 41.8 μL were combined with 11 μL of 10× Blocking agent and 2.2 μL 25× Fragmentation buffer, and incubated at 60 °C for 30 minutes.
|
| |
|
| Hybridization protocol |
The fragmented cRNA was then mixed with 55 μL 2× GEx Hybridization buffer. 100 μL of the mixture was used to the Agilent Rat Whole Genome 4×44K Array Chip (Agilent Technologies), and hybridized at 65 °C for 17 hours in an Agilent Microarray Hybridization Chamber (Agilent, G2534A) in rotating Hybridization Oven (Agilent, G2545A). After hybridization, array slides were washed with Agilent Gene expression Wash Buffer I for 1 minute at room temperature and followed by a 1 min wash with Agilent Gene expression Wash Buffer II (Agilent, 5188-5327) at 37 °C. Slides were finally rinsed with acetonitrile for cleaning up and drying.
|
| Scan protocol |
Hybridized arrays were scanned at 5 μm resolution on a Genepix 4000B Microarray Scanner (Axon Instruments, Sunnyvale, CA).
|
| Description |
Gene expression after dietary high protein diet for six weeks in rat colon
|
| Data processing |
Expression data were normalized through quantile normalization using the Robust Multichip Average algorithm. Array data were normalized to the global median signal intensity value and values were log base 2 transformed. Low-intensity and unreliable results were removed using a filter on flags feature, with standardized software algorithms
|
| |
|
| Submission date |
Feb 11, 2015 |
| Last update date |
Apr 05, 2016 |
| Contact name |
Weiyun Zhu |
| Organization name |
Nanjing Agricultural University
|
| Department |
College of Animal Science and Technology
|
| Lab |
Laboratory of Gastrointestinal Microbiology
|
| Street address |
No.1, Weigang, Xuanwu District
|
| City |
Nanjing |
| State/province |
Jiangsu |
| ZIP/Postal code |
210095 |
| Country |
China |
| |
|
| Platform ID |
GPL14746 |
| Series (1) |
| GSE65862 |
Transcriptomic response in colon of rat with increased dietary protein content |
|
