|
| Status |
Public on Jul 13, 2007 |
| Title |
0 hr Reference, biological rep 3 |
| Sample type |
RNA |
| |
|
| Source name |
All living Caenorhabditis elegans sorted cells from freshly dissociated embryos
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
embryonic cells from wildtype (N2 isolate) nematodes
|
| Growth protocol |
Gravid adults expressing myo-3::GFP were subjected to bleach/NaOH treatment to release embryos. Embryos were harvested and then treated with chitinase to degrade the eggshell. Cells were dissociated and either sorted immediately (0hr dataset) or plated on poly-L-lysine coated dishes for 24 hrs to allow differentiation and then removed from the culture dish and sorted (24 hr dataset). Cells were sorted using the FACStar Plus (Becton Dickinson, San Jose, CA) that had been flushed with egg buffer. ~90% enrichment of GFP+ cells was obtained. For Reference dataset, wildtype (N2) animals were subjected to the same dissociation/cell culture treatment. FACS isolated all non-propidium iodide stained (i.e. living) cells.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from sorted cells using the micro-RNA isolation kit (Stratagene) using the recommended volumes for 1 million cells.
|
| Label |
biotin
|
| Label protocol |
A 2-round IVT protocol (modified from the Affymetrix small-sample protocol) was used to convert 100 ng total RNA into biotinylated cRNA.
|
| |
|
| Hybridization protocol |
Following fragmentation, 10-15 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Caenorhabditis elegans Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the Affymetrix GeneChip Scanner 3000 7G.
|
| Description |
Gene expression data from all freshly dissociated wildtype embryonic nematode cells
|
| Data processing |
Data were normalized using Robust Multiarray analysis (RMA) in GeneTraffic (Stratagene). RMA normalized data were subjected to Significance Analysis of Microarray (SAM, Stanford) analysis. Enriched genes were obtained by finding genes with a 1.7x enriched at a False Discovery Rate (FDR) of less than or equal to 1.8% (0hr) or less than or equal to 1.2% (24hr)..
|
| |
|
| Submission date |
Jul 12, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
David Miller |
| E-mail(s) |
david.miller@vanderbilt.edu
|
| Phone |
6153433447
|
| Fax |
6159365673
|
| URL |
http://exploration.vanderbilt.edu/news/news_worm.htm
|
| Organization name |
Vanderbilt University
|
| Department |
Cell and Developmental Biology
|
| Street address |
465 21st Avenue South
|
| City |
Nashville |
| State/province |
TN |
| ZIP/Postal code |
37232-8240 |
| Country |
USA |
| |
|
| Platform ID |
GPL200 |
| Series (1) |
| GSE8462 |
The embryonic muscle transcriptome of Caenorhabditis elegans |
|
