|
| Status |
Public on Sep 01, 2009 |
| Title |
TAg transgenic enterocytes, biological rep1 |
| Sample type |
RNA |
| |
|
| Source name |
TAg transgenic villi enterocytes
|
| Organism |
Mus musculus |
| Characteristics |
age: 2.4 months
gender: Male
cell type: TAg transgenic enterocytes
biological replicate: 1
|
| Treatment protocol |
The intestine was removed in its entirety from the abdominal cavity of mice, from the exit of the stomach to the entry into the large intestine. The intestines were then opened along their cephalocaudal axis, washed with phosphate-buffered saline, and rolled from the duodenum to the ileum. Rolls from wild-type, TAg and TAg mutant transgenic mice were then processed to isolate villi cell types by LCM. Briefly, the rolls were quickly frozen and stored at -80C. Frozen sections were cut and immediately dehydrated in graded alcohol solutions. The dried sections were examined microscopically and the top 2/3 of 50 villous segments from each section were collected using a laser capture microscope (Pix Cell II, Arcturus, Mt. View, CA).
|
| Growth protocol |
The mice were held in our selected pathogen-free colony according to NIH guidelines for use of animals in research. The mice had access to food and water ad libitum, 24 hours a day.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from villi enterocytes with the PicoPure kit (Arcturus) and amplified twice with the RiboAmp kit (Arcturus).
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 3 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
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| |
|
| Hybridization protocol |
Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Mouse Genome 430 2.0 Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the Agilent GeneArray 3000 with 7G upgrade.
|
| Description |
TAg transgenic enterocytes, biological rep1
|
| Data processing |
The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 150. RMA was performed using BRB-Array Tools (Rich Simon, National Cancer Institute, http://linus.nci.nih.gov/BRB-ArrayTools.html).
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| |
|
| Submission date |
Jun 02, 2009 |
| Last update date |
Jun 03, 2009 |
| Contact name |
Daniel Clark |
| Organization name |
University of Pittsburgh
|
| Street address |
3501 Terrace St.
|
| City |
Pittsburgh |
| ZIP/Postal code |
15213 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE16389 |
Global analysis of gene expression by SV40 T antigen in the mouse small intestine epithelium |
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