|
| Status |
Public on Mar 25, 2008 |
| Title |
E6.0 embryo ratio to E2.5 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
mRNA from mouse embryo E6.0
|
| Organism |
Mus musculus |
| Characteristics |
ICR timed mated
|
| Treatment protocol |
The embryos were not chemically treated in any way
|
| Growth protocol |
ICR embryos were collected by flushing the oviduct for stages 2.5, or the uterus for stages 3.5. Stages 4.5 and older were dissected from the uterus. None of the vaginal plug was considered as E0.5.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Embryos from the different stages were pooled for mRNA extraction with the Micro-Fast Track isolation kit (Invitrogen, 45-0036). Three independent pools (for three independant experiments) were made for each stage, in average 106 embryos at E2.5, 61 at E3.5, 47 at E4.5, 28 at E4.8, 17 at E5.5 and 19 at E6.5.
|
| Label |
Cy3
|
| Label protocol |
SMART (Clontech) reverse transcription and PCR were adapted for each developmental stage to amplify cDNAs. To generate probes for array hybridization, 1 microgram of cDNA was labeled by incorporation of either Cy5 or Cy3-dCTP during random hexamer-primed primer extension in the presence of Klenow DNA polymerase (Roche) according to Livesey et al. (2000)
|
| |
|
| Channel 2 |
| Source name |
mRNA from mouse embryo E2.5
|
| Organism |
Mus musculus |
| Characteristics |
ICR timed mated
|
| Treatment protocol |
The embryos were not chemically treated in any way
|
| Growth protocol |
ICR embryos were collected by flushing the oviduct for stages 2.5, or the uterus for stages 3.5. Stages 4.5 and older were dissected from the uterus. None of the vaginal plug was considered as E0.5.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Embryos from the different stages were pooled for mRNA extraction with the Micro-Fast Track isolation kit (Invitrogen, 45-0036). Three independent pools (for three independant experiments) were made for each stage, in average 106 embryos at E2.5, 61 at E3.5, 47 at E4.5, 28 at E4.8, 17 at E5.5 and 19 at E6.5.
|
| Label |
Cy5
|
| Label protocol |
SMART (Clontech) reverse transcription and PCR were adapted for each developmental stage to amplify cDNAs. To generate probes for array hybridization, 1 microgram of cDNA was labeled by incorporation of either Cy5 or Cy3-dCTP during random hexamer-primed primer extension in the presence of Klenow DNA polymerase (Roche) according to Livesey et al. (2000)
|
| |
|
| |
| Hybridization protocol |
Labelled probes were hybridised according to Wigle et al. (2002).
|
| Scan protocol |
Slides were scanned with a Genepix Axon 4000 microarray scanner.
|
| Description |
mouse embryos at different stages to identify genes during the development of ICM, TE, and PrE
|
| Data processing |
Spot intensities were quantified and median back ground corrected with the supplied Genepix software and exported as tables. Duplicate samples were analyzed and probe spot intensities were averaged. Expression data was set as a log2 ratio of the genes expression relative to E2.5.
|
| |
|
| Submission date |
Jun 29, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Brian Joseph Cox |
| E-mail(s) |
b.cox@utoronto.ca
|
| Organization name |
University of Toronto
|
| Department |
Physiology
|
| Lab |
Cox System Biology
|
| Street address |
1 King's College Circle, Rm 3360
|
| City |
Toronto |
| State/province |
Ontario |
| ZIP/Postal code |
M5S1A8 |
| Country |
Canada |
| |
|
| Platform ID |
GPL519 |
| Series (1) |
| GSE8339 |
Early mouse embryo development |
|
