|
| Status |
Public on Aug 06, 2014 |
| Title |
laevis_DBM_s28_2 |
| Sample type |
RNA |
| |
|
| Source name |
Whole embryo, DBM-injected, stage 28
|
| Organism |
Xenopus laevis |
| Characteristics |
Stage: 28
injected construct: DBM
|
| Treatment protocol |
Embryos were injected into one blastomere at the two cell stage with 1.5 ng of one of the following: capped RNA constructs synthesized in vitro; a DNA Binding mutant of suppressor of hairless (DBM), a construct that suppresses Notch signaling; the Intracelluar Domain (ICD), which activates the Notch signaling pathway; or Green Fluorescent Protein (GFP) as a tracer and control for the injection procedure. The DBM and ICD constructs were kind gifts from Dr. Chris Kintner. Capped RNA was synthesized in vitro using mMessage Machine (Ambion) following the manufacturer’s protocol.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Embryos were raised to stages 18, 28, and 38 for RNA collection. To obtain total RNA, 10 embryos from each stage and condition were homoginized in Tri Reagent and extracted with BCP according to the manufacturer's protocol. RNA from the aqueous phase was purified using the Qiagen RNeasy Mini kit.
|
| Label |
biotin
|
| Label protocol |
500 ng of RNA from each sample was used as input. The 3’ IVT Express kit was used to prepare the samples according to the manufacturer’s instructions with the exception that the in vitro transcription reaction was carried out for 16 hours rather than the 4 hours recommended by the manufacturer.
|
| |
|
| Hybridization protocol |
500 ng of RNA from each sample was used as input. The 3’ IVT Express kit was used to prepare the samples according to the manufacturer’s instructions with the exception that the in vitro transcription reaction was carried out for 16 hours rather than the 4 hours recommended by the manufacturer.
|
| Scan protocol |
Scanning was performed using a GeneChip Scanner 3000 7G according to manufacturer's protocols.
|
| Description |
Downregulation of the notch pathway during tailbud formation
|
| Data processing |
Differentially expressed (DE) genes were determined using a two-sample linear design model with group-means parameterization with Bayes correction according to the method of Smyth. This analysis was implemented with the limma packages in Bioconductor and reported with p-values adjusted for multiple tests using the Benjamini-Hochberg procedure. Normalized hybridization intensities are reported in the data table as log2 transformed values. Normalized values before log2 transformation were used for statistical testing.
|
| |
|
| Submission date |
Aug 05, 2014 |
| Last update date |
Aug 06, 2014 |
| Contact name |
Margaret Saha |
| E-mail(s) |
mssaha@wm.edu
|
| Phone |
7572212407
|
| Organization name |
College of William and Mary
|
| Department |
Biology
|
| Street address |
540 Landrum Drive
|
| City |
Willimsburg |
| State/province |
VA |
| ZIP/Postal code |
23187 |
| Country |
USA |
| |
|
| Platform ID |
GPL10756 |
| Series (1) |
| GSE60127 |
Perturbation of the Notch pathway in Xenopus laevis embryos |
|
