|
| Status |
Public on May 31, 2010 |
| Title |
Caenorhabditis elegans grown with sufficient Biotin 1 |
| Sample type |
RNA |
| |
|
| Source name |
Caenorhabditis elegans in grown in minimal media with conditions of biotin sufficiency
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
strain: N2
developmental stage: L4 stage
environmental factor: biotin suffficient diet
|
| Treatment protocol |
Caenorhabditis elegans were collected and resuspended in PBS pH 7.0 buffer.
|
| Growth protocol |
Biotin deficiency for C. elegans strain N2 was achieved by feeding nematodes synchronized in L1 stage with an overnight culture of E.coli biotin auxotrophic (MG1655bioB:kan, kindly donated by Dr. Cronan) grown in minimal medium (MM) with 5 and 0.05 nM of biotin (sufficient and deficient, respectively). To generate deficiency in worms, avidin (2U/ml) was added at each condition. Worms were allowed to grow until the L4 stage.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted was extracted with the TRIzol® method (Invitrogen) according to the manufacturer’s instructions. RNA was subsequently loaded onto RNA-binding columns from QIAGEN’s RNeasy kit (Qiagen, Valencia, CA) to remove contamination with genomic DNA and proteins before further processing.
|
| Label |
biotin
|
| Label protocol |
Total RNA (5 ug) was reverse transcribed using One Cycle cDNA Synthesis kit (Affymetrix, Santa Clara, CA) and the obtained double-stranded cDNA was purified with GeneChip Sample Cleanup Module (Affymetrix, Santa Clara, CA). The obtained cDNA was used as a template for in vitro transcription using GeneChip IVT Labeling Kit (Affymetrix, Santa Clara, CA).
|
| |
|
| Hybridization protocol |
Following fragmentation, 15 ug of cRNA were hybridized for 16 hr at 45C with rotation (60 rpm) on the array (Affymetrix, Santa Clara, CA). GeneChips were washed and stained using Affymetrix fluidics station 450 with streptavidin-R-phycoerythrin, and the signal was amplified using a biotinylated goat anti-streptavidin antibody followed by another SAPE staining (Hybridization, Washing and Staining Kit, Affymetrix, Santa Clara, CA).
|
| Scan protocol |
Microarrays were scanned using Affymetrix Gene Array Scanner 3000 7G Plus (Affymetrix, Santa Clara, CA).
|
| Description |
Gene expression data from Caenorhabditis elegans grown with sufficient biotin
|
| Data processing |
The data were analyzed with Affymetrix GCOS 1.4 software by comparing means of signals intensity from RNA samples derived from biotin sufficient and deficient organisms.
|
| |
|
| Submission date |
Sep 17, 2009 |
| Last update date |
Dec 23, 2009 |
| Contact name |
Pablo Antonio Velázquez-Arellano |
| E-mail(s) |
velare@servidor.unam.mx
|
| Phone |
(52) 55-5622-6421
|
| Fax |
(52) 55-5606-3489
|
| Organization name |
Institute for Biomedical Research, UNAM
|
| Department |
Molecular Biology and Biotechnology
|
| Street address |
Av. IMAN No. 1, 4th floor
|
| City |
MEXICO, D.F. |
| ZIP/Postal code |
04530 |
| Country |
Mexico |
| |
|
| Platform ID |
GPL200 |
| Series (1) |
| GSE18157 |
Gene expression underlying the effects of biotin deficiency in rat liver, worms and yeast |
|
