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| Status |
Public on Mar 23, 2021 |
| Title |
Ts_GC_non salt_rep_4_Light |
| Sample type |
RNA |
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| Source name |
Guard cells
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| Organism |
Eutrema salsugineum |
| Characteristics |
age: 9 weeks
|
| Treatment protocol |
Two different salt treatment protocols, a short- and long-term, were applied to 7 weeks old Thellungiella plants by soaking the soil pots in salt solutions each time for two hours. In the long-term salt stress experiment plants were treated three times : (i) On day 1 and 2 (75 mM NaCl), (ii) on day 7 and 8 (150 mM NaCl), and (iii) on day 13 and 14 (200 mM NaCl) named as 3x salt. In the short-term salt stress experiment plants were treated with 200mM NaCl on day 13 and 14 (named as 1x salt) while the control group faced tap water (named as (-) salt). All samples were collected at the end of the salt experiments on day 15 to ensure that plants have the same plant age and developmental stage.
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| Growth protocol |
Thellngiella salsuginea (accession Sahandong) plants were grown in pots (7 cm diameter, 200 ml volume) filled with 91 ± 7 g soil (Einheitserde Typ P, Germany). Plants were illuminated for 12 hours (10 A.M. to 10 P.M.) with a light intensity of 80-110 μmol s-1 m-2 using fluorescent lamps (Osram, L58W/77 FLUORA; Munich, Germany) at 22°C in light and 16°C in dark and 60% relative air humidity.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the NucleoSpin RNA kit (Macherey-Nagel, Gmbh& co. Germany) according to manufacturer’s protocol from salt (1x and 3x salt) and non-salt treated plants kept in light for four hours after beginning of the light period (from 10 am to 2 pm). Four guard cell samples of each treatment were prepared and hence, a total 12 samples for Thellungiella were used for microarray analysis.
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| Label |
Cy3
|
| Label protocol |
For labeling, the Low Input QuickAmp Labeling Kit (Agilent Technologies) was used to generate fuorescent cRNA (complementary RNA). For 1st strand synthesis, either oligo-dT primer or random primer / oligo-dT primer mixture (WT primer) was used. After 2nd strand synthesis, an in vitro transcription for synthesis of cRNA labelled with cyanine 3-CTP was performed
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| |
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| Hybridization protocol |
For hybridization, the Agilent Gene Expression Hybridization Kit (Agilent Technologies) was used. 600 ng of each cRNA was hybridized on 8x60K microarray at 65°C for 17
|
| Scan protocol |
Fluorescence signals on microarrays were detected by the SureScan Microarray Scanner (Agilent Technologies) at a resolution of 3 micron for SurePrint G3 Gene Expression Microarrays and 5 micron for HD Microarray formats, generating a 20 bit TIFF file.
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| Description |
Gene expression after non salt treatment
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| Data processing |
Agilent's Feature Extraction software version 11 was used to read and process the TIFF files. Data pre-processing was performed using the Bioconductor software (Huber et al. 2015) with the statistical programming environment R (R Development Core Team, 2011). Normalization has been performed using negative control probes and quantile normalization using negative and positive control probes as implemented in the neqc function (Shi et al. 2010) of the Limma package (Ritchie et al. 2015).
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| |
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| Submission date |
Mar 22, 2021 |
| Last update date |
Mar 23, 2021 |
| Contact name |
Sohail Mehmood Karimi |
| E-mail(s) |
suhailmkarimi@gmail.com
|
| Organization name |
University of Wuerzburg
|
| Department |
Botany 1
|
| Lab |
Rosalia Deeken
|
| Street address |
Julius-von-Sachs-Platz 2,
|
| City |
Wuerzburg |
| State/province |
Bavaria |
| ZIP/Postal code |
97082 |
| Country |
Germany |
| |
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| Platform ID |
GPL19319 |
| Series (1) |
| GSE169311 |
Whole genome transcriptome profiling of Thellungiella salsuginea guard cells under saline growth conditions |
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