|
| Status |
Public on Mar 23, 2010 |
| Title |
Tie2_E15.5_1 |
| Sample type |
RNA |
| |
|
| Source name |
small blood vessels
|
| Organism |
Mus musculus |
| Characteristics |
gene reported: Tg(TIE2GFP)287Sato / MGI:3056937
strain: CD-1
developmental stage: 15.5 dpc
theiler stage: TS23
|
| Treatment protocol |
pool size: 4-5 Embryonic Kidneys
Pooled sample: Yes
Dissection Method: trypsinization & fluorescent activated cell sorting
Tie2-GFP transgenic mice are time-mated. Pregnant Tie2-GFP transgenic mice are euthanized by standard carbon dioxide asphyxiation. All fetuses are killed by decapitation with a scalpel. Fetal kidneys are dissected from fetuses and placed in ice-cold PBS. Embryonic kidneys are incubated in the presence of 300µl of trypsin for 5 minutes at 37 ºC. Kidneys are then dissociated by titurating in the presence of 600µl of ice-cold 10%FBS/PBS. The kidneys are pelleted at 5000 rpm, 4 ºC for 5 minutes. The media is aspirates and the cell pellet is resuspended in 200µl of ice-cold 2%FBS/PBS. Filter cells through 70 micron mesh filter and proceed with FACS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Potter protocol: 'RNA purification'
|
| Label |
Biotin
|
| Label protocol |
The FL-Ovation cDNA Biotin Module V2 (NuGEN) is used to chemically modify fragments and label target sense cDNA.
|
| |
|
| Hybridization protocol |
Affymetrix standard protocol
Amount labeled target hybridization to array: 2.5µg
The arrays were washed and and stained using a Fluidics Station 450 (Affymetrix) utilizing the fluidics protocol FS450-0007.
|
| Scan protocol |
Probe arrays were scanned using an Affymetrix GeneChip Scanner 3000 7G and Affymetrix scanning software Genechip Operating Software Version 1.4.
|
| Description |
>> Amplification protocol <<
Target Amplified manufacturer/kit: WT-Ovation Pico RNA Amplification System (NuGEN)
Target Amplified protocol: Potter protocols
Rounds of amplification: 1
URL: http://www.gudmap.org/gudmap/pages/mic_submission.html?id=GUDMAP:13556
|
| Data processing |
Analysis method: Affymetrix Expression Console and GeneSpring 10.
|
| |
|
| Submission date |
Mar 22, 2010 |
| Last update date |
Dec 30, 2010 |
| Contact name |
GUDMAP Developers |
| E-mail(s) |
[email protected]
|
| Phone |
+44 131 651 8500
|
| Organization name |
IGMM MRC Human Genetics Unit
|
| Lab |
GUDMAP Database Group
|
| Street address |
Crewe Road
|
| City |
Edinburgh |
| ZIP/Postal code |
EH4 2XU |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL6246 |
| Series (1) |
| GSE20991 |
Gene expression profiles of E15.5 endothelial cells isolated from TIE2-GFP transgenic mice using FACS. (GUDMAP Series ID:38) |
|
