|
| Status |
Public on Aug 01, 2011 |
| Title |
0minSucChemostat_30minMeOH_BioRep2_TechRep1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Succinate limited chemostat grown cells
|
| Organism |
Methylorubrum extorquens AM1 |
| Characteristics |
strain: AM1
|
| Treatment protocol |
Cells were processed as previously described ( Okubo et al., OMICS. 2007, 11:325-340.).
|
| Growth protocol |
For a starting point (time = 0 min), the carbon-shift experiment utilized cells from a steady-state chemostat culture of M. extorquens AM1 growing on limiting succinate. M. extorquens AM1 was cultivated in a minimal medium (Okubo and Skovran et al, 2007) containing 3.7 mM succinate as a growth-limiting nutrient and 50 mg/mL rifamycin in a 2.2-liter bench-top BioFlo 110 Modular Fermentor (New Brunswick Scientific) with a 1 liter working volume, resulting in an OD600 of ~0.63. Dilution rate was maintained at 0.163 h-1. Cells were maintained at 28 degrees and at pH 6.8 as described previously (Guo and Lidstrom, 2006). After the culture reached steady-state, cells were transferred to flasks before 50 mM methanol addition and incubated at 28 degrees C shaking in a floor incubator, harvesting cells/RNA at time= 0, 10, 30, 60, 120, 240 and 360 min post methanol addition. For a carbon starvation control, cells were transferred to a flask and incubated in a separate incubator (to avoid methanol vapours) as described above but with no methanol added.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated as described by Okubo and Skovran et al., OMICS. 2007, 11:325-340.
|
| Label |
A555
|
| Label protocol |
cDNA production and labelling were carried out by MOgene (St. Louis, MO).
|
| |
|
| Channel 2 |
| Source name |
T=30 min post MeOH addition
|
| Organism |
Methylorubrum extorquens AM1 |
| Characteristics |
strain: AM1
|
| Treatment protocol |
Cells were processed as previously described ( Okubo et al., OMICS. 2007, 11:325-340.).
|
| Growth protocol |
For a starting point (time = 0 min), the carbon-shift experiment utilized cells from a steady-state chemostat culture of M. extorquens AM1 growing on limiting succinate. M. extorquens AM1 was cultivated in a minimal medium (Okubo and Skovran et al, 2007) containing 3.7 mM succinate as a growth-limiting nutrient and 50 mg/mL rifamycin in a 2.2-liter bench-top BioFlo 110 Modular Fermentor (New Brunswick Scientific) with a 1 liter working volume, resulting in an OD600 of ~0.63. Dilution rate was maintained at 0.163 h-1. Cells were maintained at 28 degrees and at pH 6.8 as described previously (Guo and Lidstrom, 2006). After the culture reached steady-state, cells were transferred to flasks before 50 mM methanol addition and incubated at 28 degrees C shaking in a floor incubator, harvesting cells/RNA at time= 0, 10, 30, 60, 120, 240 and 360 min post methanol addition. For a carbon starvation control, cells were transferred to a flask and incubated in a separate incubator (to avoid methanol vapours) as described above but with no methanol added.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated as described by Okubo and Skovran et al., OMICS. 2007, 11:325-340.
|
| Label |
A647
|
| Label protocol |
cDNA production and labelling were carried out by MOgene (St. Louis, MO).
|
| |
|
| |
| Hybridization protocol |
Hybridization was carried out by MOgene.
|
| Scan protocol |
Scanning was carried out by MOgene.
|
| Description |
S17_0vs30_2a
|
| Data processing |
Data processing was carried out as described in Okubo and Skovran et al, 2007. Essentially, data from duplicated probes within each array were averaged and the two technical replicates for each biological replicate were merged together by averaging the Log10 ratios of set "a" with the inverse Log10 ratios of set "b" since the replicates were dye-swaps (see GSE22031 supplementary file: Processed_Matrix.xls). Biological replicates were not averaged and are displayed as "Set 1" and Set 2" in published Supplementary Tables.
|
| |
|
| Submission date |
May 27, 2010 |
| Last update date |
Aug 01, 2011 |
| Contact name |
Elizabeth Skovran |
| E-mail(s) |
[email protected]
|
| Fax |
206-616-5721
|
| Organization name |
University of Washington
|
| Department |
Chemical Engineering
|
| Lab |
Mary Lidstrom
|
| Street address |
616 NE Northlake Place, Benjamin Hall Rm 440
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98105 |
| Country |
USA |
| |
|
| Platform ID |
GPL6262 |
| Series (1) |
| GSE22031 |
Transition from succinate to methanol growth in Methylobacterium Extorquens AM1 |
|
