|
| Status |
Public on Mar 12, 2007 |
| Title |
small intestine EPI-MES Epi C |
| Sample type |
RNA |
| |
|
| Source name |
mouse small intestine E18.5 days epithelium C
|
| Organism |
Mus musculus |
| Characteristics |
Mouse small intesine (E18.5) was separated to epithelium and mesenchyme. This sample is EPI C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Tissue separation was done as previously described. Briefly, E18.5 embryos from C57BL/6 females were removed from the yolk sac and amnion and placed into ice-cold PBS containing penicillin/streptomycin. The entire small intestine, from duodenum to cecum, was isolated, cut open longitudinally, placed into 1.0 ml of cold Cell Recovery Solution (BD Biosciences) in a 12-well plate and incubated for nine hours at 4°C. On ice, each plate was agitated by hand for 30 to 45 minutes until all epithelium sloughed off, as determined by microscopic examination. The mesenchyme was gently removed from the more delicate epithelial fragments with sterile forceps and rinsed in a dish of sterile PBS to remove any remaining epithelial cells. Mesenchyme from three embryos was pooled into a 15 ml conical tube then snap frozen in liquid nitrogen; six such pooled samples were prepared. The remaining epithelial tissue was washed twice with 10 ml of cold PBS and collected by gentle centrifugation (100xg for 7 minutes at 4°C). Intestinal epithelium from three embryos was pooled into a 15 ml conical tube then snap frozen in liquid nitrogen; six pooled samples were prepared. Each pool of tissue was subsequently homogenized in 1.0 mls TRIzol (Invitrogen), on ice, and RNA prepared per the TRIzol protocol. Total RNA was further purified using the RNeasy Mini Kit (Qiagen), and quality assessed by electrophoresis of 2.0 µg RNA on a 1% agarose gel. To assess epithelial vs. mesenchymal purity of RNA, we performed RT-PCR for both epithelial-specific mRNAs (Vil1 and Prkg2) and mesenchymal-specific mRNAs (Vim, Madcam1, and Actg2). Three epithelial and three mesenchymal RNA fractions exhibiting no or minimal contamination with mesenchymal and epithelial mRNAs, respectively, were selected from among the samples for Affymetrix microarray analysis.
|
| Label |
biotin
|
| Label protocol |
samples were labelled according to affymetrix protocols:
http://www.affymetrix.com/support/technical/manual/expression_manual.affx
|
| |
|
| Hybridization protocol |
samples were bybridized according to affymetrix protocols:
http://www.affymetrix.com/support/technical/manual/expression_manual.affx
|
| Scan protocol |
microarray was scanned according to affymetrix protocols.
|
| Description |
There are 6 chips in the microarray experiment. 3 for epithilium of mouse small intestine; 3 for mesenchyme of mouse small intestine.
|
| Data processing |
Microarray CEL files was processedusing RMA to get gene expression values
|
| |
|
| Submission date |
Nov 27, 2006 |
| Last update date |
Aug 28, 2018 |
| Contact name |
Xing Li |
| Phone |
734-647-0171
|
| URL |
http://0-www-ncbi-nlm-nih-gov.brum.beds.ac.uk/entrez/query.fcgi?cmd=retrieve&db=pubmed&list_uids=17299133&dopt=Abstract
|
| Organization name |
Universtiy of Michigan
|
| Department |
Bioinformatics Program & CDB
|
| Lab |
Dr. Gumucio Lab
|
| Street address |
109 Zina Pitcher Place,045 BSRB
|
| City |
Ann Arbor |
| State/province |
MI |
| ZIP/Postal code |
48109-2200 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE6383 |
Mouse small intestine epithelium vs. mesenchyme |
|
| Relations |
| Reanalyzed by |
GSE119085 |
