|
| Status |
Public on Jun 30, 2015 |
| Title |
Xoo LPS rep1 |
| Sample type |
RNA |
| |
|
| Source name |
Rice leaf midvein 15 h after treatment with Xoo LPS
|
| Organism |
Oryza sativa |
| Characteristics |
treatment: LPS extracted from Xoo BXO43
tissue: Midvein of rice leaf
|
| Treatment protocol |
Leaves from plants in a single pot were either injected with 200 micro liter Xoo LPS (100 µg/ml) or 200 µl control solution. For each treatment, injections were performed into the mid veins of 5 leaves. 15 h after injection, 4 cm long portions of mid vein was cut out from below the point of injection and snap freezed in liquid nitrogen. Each such pot constituted one biological replicate.
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| Growth protocol |
Rice seeds were germinated in petriplates containing moist bloting paper. Seedlings were later transferd to soil in pots. One week old seedlings were transplanted to a field. 30 days old rice plants were again transferred to pots and kept in green house conditions for 10 days prior to the experimental treatments.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using the TrizolTM (Life technologies, California, U. S. A) reagent using the manufacturer's protocol. Mid veinal tissues of rice leaves were isolated, frozen in liquid nitrogen and subsequently used for sample preparation.
|
| Label |
Biotin
|
| Label protocol |
250 ng of RNA was used as the starting material in cDNA preparation. First strand cDNA synthesis was done using the components of GeneChip IVT express kit (Affymetrix, California, U. S. A.) using the manufacturer’s protocol. 30 µl of in vitro transcription (IVT) master mix containing IVT biotin labeled uridine trinucleotide, IVT labeling buffer and IVT enzyme mix was mixed thoroughly with 30 μl of double stranded cDNA sample and incubated at 40˚C for 4 hours to obtain antisense RNA (aRNA).
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| |
|
| Hybridization protocol |
The hybridization of fragmented and labeled aRNA with the rice GeneChip array (Affymetrix, California, U. S. A.) was done according to the manufacturer’s protocol at 45˚C for 16 hours in an Affymetrix Hybridization Oven 640. The GeneChips were later washed and stained using the Affymetrix GeneChip Fluidics station 450 according to the manufacturer’s protocol.
|
| Scan protocol |
The stained GeneChips were later scanned using the Affymetrix GeneChip Scanner 450 to obtain the DAT file which contains the optical image of the hybridized chip in TIFF format
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| Description |
Gene expression data from rice leaf treated with lipopolysaccharide isolated from Xanthomonas oryzae pv. oryzae
|
| Data processing |
The DAT file was then processed using the Affymetrix Genechip Command Console (GCOS) to obtain the CEL files which contain the light intensity information corresponding to each probe location in the GeneChip. CEL files were further processed along with the CDF (Chip description file) to obtain the TXT file which contains the intensity values of each probeset. The data were normalized using the probe intensity logarithmic error (PLIER) algorithm.
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| |
|
| Submission date |
Sep 29, 2014 |
| Last update date |
Jun 30, 2015 |
| Contact name |
Ramesh V. Sonti |
| E-mail(s) |
sonti@ccmb.res.in
|
| Organization name |
Centre for cellular and molecular biology (CCMB)
|
| Department |
Plant Microbe Interactions
|
| Street address |
Uppal Road
|
| City |
Hyderabad |
| State/province |
Andhra Pradesh |
| ZIP/Postal code |
500007 |
| Country |
India |
| |
|
| Platform ID |
GPL2025 |
| Series (1) |
| GSE61832 |
Gene expression changes induced in Rice (Oryza sativa) leaves by lipopolysaccharide (LPS) extracted from Xanthomonas oryzae pv. oryzae (Xoo) strain BXO43 |
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