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MB Sample ID: SA035017
| Local Sample ID: | CD06241621 |
| Subject ID: | SU000644 |
| Subject Type: | Photosynthetic organism |
| Subject Species: | Chlamydomonas reinhardtii |
| Taxonomy ID: | 3055 |
| Genotype Strain: | CC125 Wild type |
| Species Group: | Microorganism |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU000644 |
| Subject Type: | Photosynthetic organism |
| Subject Species: | Chlamydomonas reinhardtii |
| Taxonomy ID: | 3055 |
| Genotype Strain: | CC125 Wild type |
| Species Group: | Microorganism |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| CD06241621 | SA035017 | FL007486 | cmp_784 | Group |
Collection:
| Collection ID: | CO000638 |
| Collection Summary: | Triplicate cultures were grown and treated as indicated and harvested at 72 h. The harvested cells were freeze-dried and weighted 10 ± 0.5 mg dry powder. To extract the metabolites, 500 uL of 75% MeOH in 0.15 M NaCl was added and metabolites were extracted using bead beating 50 Hz for 5 min. The homogenate was sonicated in water bath for 5 min and then 300 uL of 0.15 M NaCl was added and then 1 mL of chloroform:methanol (1:2) with 0.01% BHT was added. Samples were homogenized and allowed to phase separate on ice for 30 mins. Finally tubes were centrifuged for 10 min to separate the polar and non-polar layer. Top polar layer was removed and an aliquot of 200 uL was used for analysis. Samples were evaporated to dryness in SpeedVac and metabolites were reconstituted in 0.03% formic acid in analytical-grade water. Debris was removed via centrifugation and the supernatant was subjected to UPLC-MS analysis. |
| Sample Type: | Photosynthetic organism |
| Collection Method: | Centrifugation from suspension culture. |
| Collection Location: | FATTTLab Department of Biochemistry, Univ of Nebraska-Lincoln |
| Storage Conditions: | After harvest, cells were kept at -80 C until extraction. |
| Collection Vials: | 2 mL Eppendorf tubes |
Treatment:
| Treatment ID: | TR000658 |
| Treatment Summary: | For compound treatment, cells from mid-log phase culture were harvested, washed once with fresh sterile TAP media and inoculated at starting density of 5 x 105 cells/mL. Thus for the current experiment 3 different treatment conditions were generated. Cells without compound treatment were negative control for the lipid accumulation and 2 compounds were used to generate treatment conditions. All compounds were used at a final concentration of 5 M (this concentration was determined using a dose-response curve). For all cultures, 250 mL Erlenmeyer flasks with rubber stopper adopted to facilitate gas exchange were used and maintained in horizontal orbital shaking growing chamber (Innova 43; New Brunswick). At the end of 72h, cells were harvested, media was removed via centrifugation and biomass was stored at -80 C until metabolite extraction. |
| Cell Media: | Tris-Acetate Phoshphate (TAP) media. |
Sample Preparation:
| Sampleprep ID: | SP000651 |
| Sampleprep Summary: | Triplicate cultures were grown and treated as indicated and harvested at 72 h. The harvested cells were freeze-dried and weighted 10 ± 0.5 mg dry powder. To extract the metabolites, 500 uL of 75% MeOH in 0.15 M NaCl was added and metabolites were extracted using bead beating 50 Hz for 5 min. The homogenate was sonicated in water bath for 5 min and then 300 uL of 0.15 M NaCl was added and then 1 mL of chloroform:methanol (1:2) with 0.01% BHT was added. Samples were homogenized and allowed to phase separate on ice for 30 mins. Finally tubes were centrifuged for 10 min to separate the polar and non-polar layer. Top polar layer was removed and an aliquot of 200 uL was used for analysis. Samples were evaporated to dryness in SpeedVac and metabolites were reconstituted in 0.03% formic acid in analytical-grade water. Debris was removed via centrifugation and the supernatant was subjected to UPLC-MS analysis |
| Extraction Method: | Bead beating |
Combined analysis:
| Analysis ID | AN000953 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Agilent 1200 |
| Column | ACE 5 C18-300 (100 x 2.1mm) |
| MS Type | ESI |
| MS instrument type | QTOF |
| MS instrument name | Waters Synapt G2 S QTOF |
| Ion Mode | POSITIVE |
| Units | intensity |
Chromatography:
| Chromatography ID: | CH000678 |
| Instrument Name: | Agilent 1200 |
| Column Name: | ACE 5 C18-300 (100 x 2.1mm) |
| Flow Rate: | 0.1 mL/min |
| Sample Injection: | 8 uL |
| Solvent A: | 100% water; 0.1% formic acid |
| Solvent B: | 100% methanol; 0.1% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS000848 |
| Analysis ID: | AN000953 |
| Instrument Name: | Waters Synapt G2 S QTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| Ion Mode: | POSITIVE |
