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MB Sample ID: SA035016
Local Sample ID: | CD06241619 |
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 |
---|---|---|---|---|
CD06241619 | SA035016 | 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 |
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 |
Instrument Type: | QTOF |
MS Type: | ESI |
Ion Mode: | POSITIVE |