|
| Status |
Public on Mar 11, 2024 |
| Title |
Ventral tissue at E12.5, biological rep 4 |
| Sample type |
RNA |
| |
|
| Source name |
Ventral tissue at E12.5
|
| Organism |
Mus musculus |
| Characteristics |
tissue: ventral aortic tissue
isolation: Tissue isolated by laser microdissection
developmental stage: E12.5
strain: C57Bl/6
|
| Treatment protocol |
Unfixed embryos were embedded in OCT medium in presence of isopentane cooled at -65°C. Twenty μm cryostat sections were then prepared and stained before laser micro-dissection. The dorsal tissue localized in between the notochord and the roof of the aorta and the ventral tissue underneath the aorta were isolated from the same embryos.
|
| Growth protocol |
Embryos were collected from pregnant C57B/6 mice at embryonic day 12.5.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Microdissected tissues were collected on appropriate caps and then proceeded for RNA extraction using Acturus Picopure RNA isolation kit. RNAs were eluted in a final volume of 13 μl and their concentrations and integrity quantified on a bioanalyzer (Agilent).
|
| Label |
biotin
|
| Label protocol |
Between 200 and 1000 pg of total RNA was reverse transcribed following the Ovation Pico V2 system (Nugen) and the resulting double strand cDNA was used for amplification based on SPIA technology. After purification according to Nugen protocol, sens target DNA was fragmented and biotin labelled using Encore Biotin Module kit (Nugen).
|
| |
|
| Hybridization protocol |
After control of fragmentation using Bioanalyzer 2100, cDNA is then hybridized to GeneChip® Mouse Gene 2.0 ST (Affymetrix) at 45°C for 17 hours.
|
| Scan protocol |
After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G.
|
| Description |
Gene expression data from ventral aortic tissue at E12.5
|
| Data processing |
The scanned images are then analyzed with Expression Console software (Affymetrix) to obtain raw data (cel files) and metrics for Quality Controls. The observations of some of these metrics and the study of the distribution of raw data show no outlier experiment. RMA normalization is performed using R with Version 21 of Entrezgene CDF brain array.
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| |
|
| Submission date |
Oct 19, 2020 |
| Last update date |
Mar 11, 2024 |
| Contact name |
Charles Durand |
| E-mail(s) |
charles.durand@sorbonne-universite.fr
|
| Phone |
+33 1 44 27 22 84
|
| Organization name |
Sorbonne University
|
| Department |
Laboratory of Developmental Biology
|
| Lab |
CNRS UMR7622
|
| Street address |
9, quai saint-Bernard
|
| City |
Paris |
| ZIP/Postal code |
75005 |
| Country |
France |
| |
|
| Platform ID |
GPL23092 |
| Series (2) |
| GSE159591 |
Molecular analysis of the subaortic hematopoietic stem cell niche [E12.5] |
| GSE159592 |
Molecular analysis of the subaortic hematopoietic stem cell niche |
|
