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MB Sample ID: SA057939
Local Sample ID: | 161116aZKsa05_2 |
Subject ID: | SU001008 |
Subject Type: | Cultured cells |
Subject Species: | Auxenochlorella protothecoides |
Taxonomy ID: | 3075 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU001008 |
Subject Type: | Cultured cells |
Subject Species: | Auxenochlorella protothecoides |
Taxonomy ID: | 3075 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
161116aZKsa05_2 | SA057939 | FL010404 | 2 - HMP | Treatment |
Collection:
Collection ID: | CO001002 |
Collection Summary: | 1 ml algae cell culture was collected, 1 ml 70% MeOH (30% dH2O) at -80 C was added to the algae sample. Tubes were spun at 12,000 rcf to pellet for 2 min at 0 C. Supernatant was decanted and pellets stored at -80 before freeze drying at -45 C. Freeze dried samples were stored at -20 C until submission for extraction. |
Collection Protocol Filename: | StudyDesignBrendanHiggins7252016.pdf |
Sample Type: | Cultured cells |
Volumeoramount Collected: | 0.11-3 mg |
Storage Conditions: | -20℃ |
Treatment:
Treatment ID: | TR001022 |
Treatment Summary: | 3 control cultures recieved no thiamine metabolites, 3 cultures received thiamine, 3 recieved HMP, and 3 were grown on residual medium from another algae species - Chlorella sorokiniana. Samples were taken daily from each of these 12 cultures over a 5 day time course so that we can observe build-up of metabolites over time. |
Treatment Protocol Filename: | StudyDesignBrendanHiggins7252016.pdf |
Sample Preparation:
Sampleprep ID: | SP001015 |
Sampleprep Summary: | 1. Check the pH of methanol (pH 7) 2. Make the extraction solution by mixing methanol, chloroform, and water in proportions of 10 : 3 : 1 3. Rinse the extraction solution for 5 min with nitrogen, making sure that the nitrogen line was flushed out of air before using it for degassing the extraction solvent solution. 4. Add one metal ball to each Eppendorf tube, close, and place in liquid nitrogen. 5. Take the Eppendorf tubes from the liquid nitrogen and place into the tube-holder of the grinder being careful to compensate for weight, maintaining equilibrium. 6. Shake for 30s at a frequency of 25 s-1 and check that the leaves have been ground into a fine powder. Repeat if necessary, submerging in liquid nitrogen first. 7. After grinding add 500μL of pre-chilled extraction solution to each tube one by one to prevent even partial thawing of the sample. Store all samples on ice while finishing this step. 8. Vortex the sample for 20s. 9. Centrifuge for 3min at 14,000 rcf using the centrifuge Eppendorf 5415 D. 10. Remove the whole supernatant into a clean Eppendorf tube. 11. Add 800μL of extraction solvent back into the tube with the pellet and metal ball. 12. Vortex the sample again for 20s. 13. Centrifuge for 3min at 14,000 rcf using the centrifuge Eppendorf 5415 D. 14. Remove the whole supernatant and combine with the previous supernatant. 15. Dry in the Labconco Centrivap cold trap concentrator to complete dryness and submit for derivitization. |
Sampleprep Protocol Filename: | SOP_Extraction_of_Plant_Samples--Chlamydomonas.pdf |
Combined analysis:
Analysis ID | AN001583 |
---|---|
Analysis type | MS |
Chromatography type | GC |
Chromatography system | Leco Pegasus III GC |
Column | Restek Rtx-5Sil (30m x 0.25mm,0.25um) |
MS Type | EI |
MS instrument type | GC-TOF |
MS instrument name | Leco Pegasus III GC TOF |
Ion Mode | POSITIVE |
Units | Counts |
Chromatography:
Chromatography ID: | CH001112 |
Methods Filename: | Data_Dictionary_Fiehn_laboratory_GCTOF_MS_primary_metabolism_10-15-2013_general.pdf |
Chromatography Comments: | Excel generated |
Instrument Name: | Leco Pegasus III GC |
Column Name: | Restek Rtx-5Sil (30m x 0.25mm,0.25um) |
Column Pressure: | 7.7 PSI (initial condition) |
Column Temperature: | 50 - 330°C |
Flow Rate: | 1 ml/min |
Injection Temperature: | 50°C ramped to 250°C by 12°C/s |
Sample Injection: | 0.5µl |
Oven Temperature: | 50°C for 1 min, then ramped at 20°C/min to 330°C, held constant for 5 min |
Transferline Temperature: | 230°C |
Washing Buffer: | Ethyl Acetate |
Sample Loop Size: | 30 m length x 0.25 mm internal diameter |
Chromatography Type: | GC |
MS:
MS ID: | MS001461 |
Analysis ID: | AN001583 |
Instrument Name: | Leco Pegasus III GC TOF |
Instrument Type: | GC-TOF |
MS Type: | EI |
Ion Mode: | POSITIVE |
Ion Source Temperature: | 250°C |
Ionization Energy: | 70eV |
Mass Accuracy: | Nominal |
Source Temperature: | 250°C |
Scan Range Moverz: | 85-500 |
Scanning Cycle: | 17 Hz |
Scanning Range: | 80-500 Da |
Skimmer Voltage: | 1850 V |