|
| Status |
Public on May 11, 2011 |
| Title |
L4 larvae, biological rep2 |
| Sample type |
RNA |
| |
|
| Source name |
C. elegans L4 larvae
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
strain: Bristol N2
age: L4
tissue: whole larva
|
| Treatment protocol |
Animals were harvested by flooding NGM plates with M9 buffer, collected into a 15 mL tube and allowed to settle. After removing most of the supernatant, animals were transferred to a cryovial and resuspended in 1 mL Trizol. After vortexing to disrupt corpses, samples were flash frozen in liquid nitrogen and stored at -80˚ C until RNA was extracted.
|
| Growth protocol |
Eggs derived from bleach-treated gravid adult C. elegans were hatched in M9 buffer and starved overnight at 20˚C to yield a developmentally synchronized population of L1 larvae. L1 larvae were then dropped to NGM (Nematode Growth Medium) plates seeded with E. coli strain OP50 and incubated at 20˚C. After 2 days, animals had reached the L4 larval stage and were split into two groups. RNA was prepared immediately from the first group, as described below, for hybridization to microarrays. The second group of L4s was transferred to OP50-seeded NGM plates containing 0.05 mg/mL FUDR (to prevent hatching of eggs) by chunking. These animals were aged at 20˚C and harvested at either day 6 or day 15 of adulthood for RNA extraction. This protocol was performed in triplicate to produce three independent biological replicates for microarray analysis.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
RNA was quantified and quality confirmed using an Agilent 2100 Bioanalyzer. 100 ng of total RNA was amplified and labeled using the NuGen Ovation RNA amplification v2 kit.
|
| |
|
| Hybridization protocol |
Samples were hybridized to GeneChip C. elegans genome microarrays at 45˚C for 16 h.
|
| Scan protocol |
GeneChips were scanned using an Affymetrix GeneChip Scanner 3000 7G.
|
| Description |
Gene expression data from a synchronized population of L4 larval stage C. elegans grown on E. coli strain OP50
|
| Data processing |
Absent/present calls were generated by analyzing the data with Microarray Suite version 5 (MAS5.0). Processed arrays were normalized and expression values output using GCRMA (Wu and Irizarry, Nature Biotechnol., 2004).
|
| |
|
| Submission date |
May 11, 2010 |
| Last update date |
May 11, 2011 |
| Contact name |
Matthew J Youngman |
| E-mail(s) |
mjy@mit.edu
|
| Phone |
617-324-3814
|
| Organization name |
MIT
|
| Department |
Biology
|
| Lab |
Dennis Kim
|
| Street address |
77 Massachusetts Ave. (68-440D)
|
| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02139 |
| Country |
USA |
| |
|
| Platform ID |
GPL200 |
| Series (1) |
| GSE21784 |
Genome-wide expression analysis during aging in C. elegans |
|
