|
| Status |
Public on Nov 05, 2011 |
| Title |
backslope rep 3 3014E B |
| Sample type |
RNA |
| |
|
| Source name |
13013E R summit/shoulder 3: top 8 cm of the topmost fully expanded leaf of four plants grown on summit/shoulder plot 3 were harvested and pooled.
|
| Organism |
Zea mays |
| Characteristics |
tissue: leaf
location: backslope
|
| Treatment protocol |
8 cm of the top most fully expanded leaf was harvested directly into liquid nitrogen and stored at -80 until RNA extraction.
|
| Growth protocol |
This research was conducted in a 65 ha corn field located in eastern South Dakota. The latitude and longitude values were 44o 10’ N and 96o 37’ W . Elevation within the field ranged from 518 to 534 m. Broadcast fertilizer was applied to this field in the fall of 2007 and spring of 2008. In the fall of 2007, 20 kg N ha-1, 23 kg P ha-1 ( 51.4 kg P2O5 ) and 36 kg K ha-1 (44.7 kg K2O) were applied. In the following spring, 102 kg N ha-1 was applied. In this field, a 97 day corn hybrid was planted in early May 2008. During the growing season, precipitation was 8.91 cm and the growing degree days from planting to silage harvest were 984. Dominant soils were the Cubden (Aeric Calciaquoll), Wauby (Aquic Hapludoll), Kranzburg (Calcic Hapludoll), and Vienna (Calic Hapludoll). Plant samples were collected on 18 July 2008. Weather conditions at time of harvest was mostly sunny with a light wind and a temperature of about 26 C. There was a thunder storm on the evening preceeding harvest.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using Trizol reagent5 and purified using Qiagen RNeasy MinElute cleanup kit6, following the manufacturer’s protocol.
|
| Label |
Alexa Fluor 555
|
| Label protocol |
First strand cDNA synthesis was performed using 1900 ng total RNA and second strand cDNA synthesis was performed using the resulting first-strand cDNA sample to make double-stranded cDNA using the Aminoallyl Message Amp II kit7. aRNA was synthesized using the resulting double-stranded cDNA. Technical replicates from each treatment were labeled with Alexa Fluor 647 or Alexa Fluor 555 dye. An Alexa Fluor 647-labeled sample from one treatment was mixed with an Alexa Fluor 555-labeled sample from another treatment.
|
| |
|
| Hybridization protocol |
46,000-element microarray chip developed by the University of Arizona was hybridized using their protocol (International Microarray Workshop Handbook, 2009; Gardiner et al. 2005).
|
| Scan protocol |
Intensities based on fluorescence for each probe were visualized and quantified with a GenePix scanner9 and GenePix Pro software.
|
| Description |
This research was conducted in a 65 ha corn field located in eastern South Dakota. The latitude and longitude values were 44o 10’ N and 96o 37’ W . Elevation within the field ranged from 518 to 534 m. Broadcast fertilizer was applied to this field in the fall of 2007 and spring of 2008. In the fall of 2007, 20 kg N ha-1, 23 kg P ha-1 ( 51.4 kg P2O5 ) and 36 kg K ha-1 (44.7 kg K2O) were applied. In the following spring, 102 kg N ha-1 was applied. In this field, a 97 day corn hybrid was planted in early May 2008. During the growing season, precipitation was 8.91 cm and the growing degree days from planting to silage harvest were 984. Dominant soils were the Cubden (Aeric Calciaquoll), Wauby (Aquic Hapludoll), Kranzburg (Calcic Hapludoll), and Vienna (Calic Hapludoll). Plant samples were collected on 18 July 2008. Weather conditions at time of harvest was mostly sunny with a light wind and a temperature of about 26 C. There was a thunder storm on the evening preceeding harvest.
Channel 2 in the raw data file
|
| Data processing |
GeneMaths XT software was used to log transform (log 2) the intensity readings and samples from each individual hybridizations were normalize against each other using default parameters in Genemaths XT software (data was treated as if it were from single channel hybridizations). Probes that had hybridization intensity less than 2 times the standard deviation plus the average of the negative controls were deleted (Horvath et al., 2007) and technical replicates for each probe were averaged to reduce any dye bias that existed. GeneMaths XT software was then used to identify P values based on ANOVA and individual t tests between treatments. Probes were considered differentially expressed if P values for any test were =0.05. Quantile normalization with centering to normalize between hybridizations.
|
| |
|
| Submission date |
Nov 04, 2011 |
| Last update date |
Nov 08, 2011 |
| Contact name |
David Horvath |
| E-mail(s) |
david.horvath@ars.usda.gov
|
| Phone |
701-239-1255
|
| Organization name |
US Dept. Agriculture
|
| Department |
Agr. Research Service
|
| Lab |
Bioscience Research Lab
|
| Street address |
1605 Albrecht Blvd
|
| City |
Fargo |
| State/province |
ND |
| ZIP/Postal code |
58105 |
| Country |
USA |
| |
|
| Platform ID |
GPL6438 |
| Series (1) |
| GSE33494 |
Drought stressed corn grown on summit/shoulder vs. well hydrated corn grown on backslope |
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