|
| Status |
Public on Oct 20, 2015 |
| Title |
PCC6803 ∆ndhD3/ndhD4/cmpA/sbtA LC 3h rep1 |
| Sample type |
RNA |
| |
|
| Source name |
cells from liquid culture grown for 3h at LC
|
| Organism |
Synechocystis sp. PCC 6803 |
| Characteristics |
strain: ∆ndhD3/ndhD4/cmpA/sbtA
|
| Treatment protocol |
For CO2-shift experiments, cells were pre-cultivated with air enriched with 5% CO2 (defined as high CO2, HC). Then, cells were harvested by centrifugation (5 min at 3000 g, 20°C). The pellet was washed and re-suspended in fresh BG11 medium of pH 7.0 at an optical density of 0.8 at 750 nm (OD750). After 1 h cultivation under HC, CO2 limitation was set by shifting the exponentially growing cultures to bubbling with ambient air containing 0.035% CO2 (defined as low CO2, LC).
|
| Growth protocol |
The glucose-tolerant strain Synechocystis sp. PCC 6803 was used as the WT (obtained from N. Murata, National Institute for Basic Biology, Okazaki, Japan). Cultures were grown in BG11 medium (Rippka et al., 1979; J Gen Microbiol 111: 1–61) buffered with 20 mM TES–KOH to pH 8.0 under conditions given before (Schwarz et al., 2011; Plant Physiol 155: 1640–1655). The ∆ndhD3/ndhD4/cmpA/sbtA mutant (= ∆4) was obtained from Prof. Teruo Ogawa (Biocenter of the University of Nagoya, Japan) and grown as WT but in presence of the corresponding, selective antibiotics.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Synechocystis 6803 cells were collected by rapid filtration on hydrophilic polyethersulfone filters (Pall Supor 800 Filter, 0.8 µm). Filters covered with cells were dissolved in 1 ml PGTX (Pinto et al., 2009; BMC Mol Biol 10: 79) and RNA was extracted as described (Hein et al., 2013; RNA Biol 10: 852–864).
|
| Label |
Cy3
|
| Label protocol |
The RNA was labeled directly, without cDNA synthesis in 2 µg aliquots with the Kreatech “ULS labeling kit for Agilent gene expression arrays” with Cy3 according to the manufacturers protocol.
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| |
|
| Hybridization protocol |
The labelled RNA was fragmented and hybridized as described by the manufacturer's instructions for Agilent one color microarrays with 1.65 µg of labeled RNA
|
| Scan protocol |
Arrays were scanned on the Agilent Technologies Scanner G2505B, using Agilent Feature Extraction Software 10.7.3.1 and the protocols GE1_107_Sep09 for Cy3 labelled arrays
|
| Data processing |
Raw data were processed with the R package Limma. Median signal intensity was quantile normalized.
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| |
|
| Submission date |
Apr 24, 2015 |
| Last update date |
Oct 20, 2015 |
| Contact name |
Stephan Klähn |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Freiburg
|
| Department |
Institute of Biology III
|
| Lab |
Genetics & Experimental Bioinformatics
|
| Street address |
Schänzlestrasse 1
|
| City |
Freiburg |
| State/province |
Baden-Würtemberg |
| ZIP/Postal code |
79104 |
| Country |
Germany |
| |
|
| Platform ID |
GPL15867 |
| Series (1) |
| GSE68250 |
Integrated functional analysis of engineered carbon limitation in the CO2/HCO3-uptake deficient ∆ndhD3/ndhD4/cmpA/sbtA mutant of Synechocystis sp. PCC 6803 |
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