|
| Status |
Public on Jun 14, 2016 |
| Title |
PCC6803_FtsHdown_+NH4_0h_iron depletion_1 |
| Sample type |
RNA |
| |
|
| Source name |
PCC6803_FtsHdown_+NH4_0h_iron depletion
|
| Organism |
Synechocystis sp. PCC 6803 |
| Characteristics |
genotype: FtsH3_down
|
| Treatment protocol |
To deplete iron, cells were transferred to iron depleted medium (residual concentration of iron about 5 µM) supplemented 10 μM deferoxamin B (DFB), an iron chelator for 24h.
|
| Growth protocol |
The previously described FtsH3down (originally termed SynFtsH3reg (Boehm et al., 2012)), FtsH2-less (ΔFtsH2) (Komenda et al., 2006), FtsH1down (Krynicka et al., 2014) strains as well as the wild type strain used for the control were derived from the non-motile glucose-tolerant Synechocystis sp. PCC 6803 strain obtained from the laboratory of Wim Vermaas. FtsH3down and its respective control (WT) were cultivated photoautotrophically in the presence of ammonium ions as described in Boehm (2012). To deplete iron, cells were transferred to iron depleted medium (residual concentration of iron about 5 µM) supplemented 10 μM deferoxamin B (DFB), an iron chelator, at an irradiance of 40 µmol photons m-2 s-1 of white light (Krynicka et al., 2014).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
WT and FtsH3down cultures were harvested by rapid filtration on hydrophilic polyethersulfone filters (Pall Supor 800 Filter, 0.8 mm). The filter covered with cells was immediately immersed in 1 ml of PGTX solution (Pinto et al., 2009) and frozen in liquid nitrogen.
|
| 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.
|
| |
|
| 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 G2505C US90900275, using Agilent Feature Extraction Software 10.7.3.1 and the protocols GE1_107_Sep09 for Cy3 labelled arrays
|
| Description |
FtsH3_down
|
| Data processing |
Raw data were processed with the R package Limma. Normexp background substraction with an offset of 50 and cyclicloess normalization was aplied to median signal intensities.
|
| |
|
| Submission date |
Jun 13, 2016 |
| Last update date |
Jun 14, 2016 |
| Contact name |
Jens Georg |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Freiburg
|
| Street address |
Schänzlestr. 1
|
| City |
Freiburg |
| ZIP/Postal code |
79104 |
| Country |
Germany |
| |
|
| Platform ID |
GPL15867 |
| Series (1) |
| GSE83303 |
The FtsH3 protease is an epistatic master regulator of the iron and phosphate stress responses in the cyanobacterium Synechocystis sp. PCC 6803 |
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