|
| Status |
Public on Aug 16, 2012 |
| Title |
W8 biological replicate 3 |
| Sample type |
RNA |
| |
|
| Source name |
third visible leaf
|
| Organism |
Zea mays |
| Characteristics |
age: 7 leaf plant
tissue: third visible leaf
line: W8
|
| Treatment protocol |
Leaf samples were cut off the plant together with leaf disc samples (from the same leaf) that were used in insect bioassays to determine mean resistance levels (see accompanying publication for details).
|
| Growth protocol |
Plants were grown in a controlled environment room at 50 ± 10% relative humidity, 14:10 light:dark cycle, with a day temperature of 24 ± 1 deg. C and a night temperature of 18 ± 1 deg. C. Lighting was supplied by alternating 1000 W sodium and halide bulbs. Plants were grown in soil mix #3 (Dowd et al. 2007, J. Ag. Food Chem. 55:3421)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from powdered tissue using Trizol as previously described (Johnson et al. 2011, Mol Gen. Genom. 285:517)
|
| Label |
Cy3
|
| Label protocol |
10 µg of total RNA was converted to cDNA with amino allyl dUTP by Superscript III reverse transcriptase. Cy5 or Cy3 dye was coupled to the modified cDNA and then purified. Details of the procedure are available in the accompanying publication.
|
| |
|
| Hybridization protocol |
40 µL of probe (40 pmol of Cy5 labeled cDNA, 40 pmol of Cy3 labeled cDNA, 25 µg of yeast tRNA, and 12 µg of salmon sperm DNA) was added to a sealed hybridization chamber containing the microarray slide. Pre-hybridization and hybridization for ~16 hours was completed as described (Hedge et al. 2000, Biotechniques 29:548) with the exception of an additional high stringency wash at room temperature.
|
| Scan protocol |
Microarray slides were scanned at 10 µm resolution with an Axon GenePix 4100A scanner using GenePix Pro software. Photomultiplier tube levels were adjusted using the Auto-PMT feature. Each image was visually scanned for aberrant signals and flagged for removal.
|
| Description |
parent 2 inbred
|
| Data processing |
For each slide, the data was normalized so that the mean of the ratio of medians of all the features was equal to one (completed using GenePix software).
Each set (one set has 4 replicates) of signal intensities (mean signal intensity minus background) for one inbred was scaled to the one replicate with the highest mean intensity. The normalized intensity values were converted to log2 values.
|
| |
|
| Submission date |
Aug 14, 2012 |
| Last update date |
Sep 12, 2012 |
| Contact name |
Eric Johnson |
| E-mail(s) |
eric.johnson2@ars.usda.gov
|
| Phone |
309-681-6177
|
| Organization name |
USDA ARS
|
| Department |
Crop Bioprotection Research
|
| Street address |
1815 N. University St.
|
| City |
Peoria |
| State/province |
IL |
| ZIP/Postal code |
61604 |
| Country |
USA |
| |
|
| Platform ID |
GPL6438 |
| Series (1) |
| GSE40107 |
Comparative transcription profiling of heritage maize lines |
|
