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| Status |
Public on Nov 30, 2010 |
| Title |
Anterior distal region of an E5.5 mouse embryo MOE430A rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Anterior distal region of an E5.5 mouse embryo
|
| Organism |
Mus musculus |
| Characteristics |
age: E5.5
tissue: Anterior distal region of embryo
genetic background: B6SJLF1×CerlP-GFP
|
| Treatment protocol |
Embyos suffered no treatment prior to the extraction procedures
|
| Growth protocol |
Embryos with fluorescently labeled AVE were collected from B6SJLF1×CerlP-GFP matings at 5.5 dpc.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Ad and Pp regions were isolated from embryos in which the AVE occupied a lateraldistal position. For the isolation of Ad regions, fragments comprising the fluorescently labeled AVE and the underlying epiblast, were microdissected. Pp regions were isolated by dissecting triangular-shaped fragments of proximal visceral endoderm and epiblast from a position diametrically opposed to the fluorescently-labeled domain. The upper limit of the Pp fragments coincided with the embryonic-extraembryonic boundary and extended in depth to roughly one third of the diameter of the egg cylinder, while its lateral limit corresponded to about one third of the height of the embryonic region. The freshly dissected fragments were immediately homogenized with TRIzol® and frozen in liquid nitrogen. The samples were then stored at -80ºC until processing.
|
| Label |
biotin
|
| Label protocol |
A modified linear amplification protocol was applied to generate the required amount of probe, according to Van Gelder et al., 1990
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| |
|
| Hybridization protocol |
15 µg of biotinylated cRNA was used in a 300-µl hybridization containing added hybridization controls. 200 µl of mixture was hybridized on Affymetrix (Santa Clara, CA, USA) GeneChip Mouse Expression Set 430 arrays for 16 h at 45°C. Standard post hybridization wash and double-stain protocols were used on an Affymetrix GeneChip Fluidics Station 400.
|
| Scan protocol |
Arrays were scanned on an Affymetrix GeneChip scanner 2500.
|
| Description |
Gene expression data from Ad region of an E5.5 mouse embryo
|
| Data processing |
Scanned arrays were analyzed first with Affymetrix MAS 5.0 software to obtain Absent/Present calls and for subsequent analysis with dChip 2010. Each set of 5 arrays (430A and 430B type) was analyzed separately from here on. Each set was normalized applying an Invariant Set Normalization Method. Then the normalized probe cell intensities of the 5 arrays were used to obtain model-based gene expression indices based on a PM-only model.
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| |
|
| Submission date |
Nov 29, 2010 |
| Last update date |
Nov 29, 2010 |
| Contact name |
Lisa Goncalves |
| E-mail(s) |
[email protected]
|
| Organization name |
CBME UALG
|
| Street address |
Campus de Gambelas, Ed8, Lab 1.17
|
| City |
Faro |
| ZIP/Postal code |
8005 |
| Country |
Portugal |
| |
|
| Platform ID |
GPL339 |
| Series (1) |
| GSE25675 |
Identification and functional analysis of novel genes expressed in the Anterior Visceral Endoderm |
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