|
| Status |
Public on Jan 26, 2007 |
| Title |
Homocysteine experiment treated 3 |
| Sample type |
RNA |
| |
|
| Source name |
CHICKEN embryo treated
|
| Organism |
Gallus gallus |
| Characteristics |
Neural tube explants were obtained from stage 9+ chicken embryos to generate neural crest cell cultures. After 4 hours in culture the cells were exposed to either media or homocysteine for 6 hours. The cultures were then processed for gene expression using the affymetrics chicken chip.
|
| Biomaterial provider |
Tom Rosenquist's Laboratory
|
| Treatment protocol |
After 4 hours, the cultures were either exposed to media or homocyteine (100uM final concentration) which has previously been shown to be teratogenic and alter neural crest cell migration, for 6 hours.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from these embryos using the RNAqueous kit from Ambion (Austin, TX) following the manufacture’s protocol. Briefly, whole embryos or tissues were placed in a guanidinium salt solution which is diluted with ethanol and passed through a glass filter cartridge which specifically binds the RNA. The RNA was then washed and recovered by a low ionic strength solution. All reagents for this protocol are included in the kit. The purity and concentration of the isolated RNA was determined by its UV-absorbance at 260 and 280 nm.
|
| Label |
Cy3;Cy5
|
| Label protocol |
Standard Affymetrics Protocol
|
| |
|
| Hybridization protocol |
Standard Affymetrics Protocol
|
| Scan protocol |
Standard Affymetrics Protocol
|
| Description |
Homocysteine Induced Alterations In Gene Expression In Neural Crest Cells
|
| Data processing |
All analyses were conducted using the BRB ArrayTool software package (http://linus.nci.nih.gov/BRB-ArrayTools.html). Pre-processing of the probe-level data was performed using the robust multiarray average (RMA) algorithm; a function of the Bioconductor Project (www.bioconductor.org/) using R-programming language (Bolstad et al, 2005; R-Development Core Team, 2004). This function performs background correction, normalization, and summarization of probe-level data. The background correction method uses the perfect match (PM) probe intensity model which is based on the assumption that the observed gene expression is the sum of both expression and background noise. Quantile-Normalization was used to normalize the PM probe intensities, and the summarization of the expression data were preformed using the Median Polish Method which fits a multichip linear model to the data and reports the expression as a log2 value (Irizarry et al., 2002). Data from both control and treated arrays were normalized as one set.
|
| |
|
| Submission date |
Jan 25, 2007 |
| Last update date |
Apr 23, 2007 |
| Contact name |
Greg Bennett |
| E-mail(s) |
gbennett@unmc.edu
|
| Phone |
402-559-3806
|
| Fax |
402-559-2873
|
| Organization name |
University of Nebraska Medical Center
|
| Department |
Genetics Cell Biology and Anatomy
|
| Street address |
Nebraska Medical Center 985805
|
| City |
Omaha |
| State/province |
NE |
| ZIP/Postal code |
68198-5805 |
| Country |
USA |
| |
|
| Platform ID |
GPL3213 |
| Series (1) |
| GSE6868 |
Homocysteine induced alterations in gene expression in neural crest cells |
|
