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MB Sample ID: SA184765
| Local Sample ID: | 71-3_13c |
| Subject ID: | SU002042 |
| Subject Type: | Plant |
| Subject Species: | Populus tremula x Populus alba |
| Taxonomy ID: | 80863 |
| Genotype Strain: | INRA-717-1B4 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002042 |
| Subject Type: | Plant |
| Subject Species: | Populus tremula x Populus alba |
| Taxonomy ID: | 80863 |
| Genotype Strain: | INRA-717-1B4 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| 71-3_13c | SA184765 | FL022717 | knockout | sample_type |
Collection:
| Collection ID: | CO002035 |
| Collection Summary: | Samples are fully expanded leaf tissue with midvein excised of three month old greenhouse grown poplar. Tissue was flash frozen and stored at -80 prior to metabolite extraction. |
| Sample Type: | Leaves |
Treatment:
| Treatment ID: | TR002054 |
| Treatment Summary: | Leaf tissue from Populus tremula x Populus alba was transformed using CRISPR-Cas9 to create functional knockouts of the UGT71L1 gene. Positive transformants were selected with Kanamycin and sequencing was conducted to confirm transformation. Clones of either wild type plants, knockouts (frame shift mutations, both alleles), intermediates (Frameshift mutation, one allele), or empty vectors (no gRNA for Cas9 construct). Plants were then grown under identical conditions for three months in a greenhouse. |
| Treatment: | Genetic |
| Plant Growth Location: | Greenhouse |
| Plant Watering Regime: | As required |
Sample Preparation:
| Sampleprep ID: | SP002048 |
| Sampleprep Summary: | Flash frozen leaf tissue was homogenized under liquid nitrogen with mortar and pestle. Tissue was then freeze dried and stored at room temperature prior to analysis. For extraction, 50 mg of freeze-dried tissue was weighed into a 2 mL cryo-vial with 4 x 5 mm steel ball bearings and 750 mL of chilled methanol. Samples were homogenized 2 x 45 seconds at 5500 rpm using a Precellys 24 Homogenizer, then sonicated for 2 minutes in a sonicating water bath. Samples were centrifuged at 12,000 rpm for 2 minutes and the supernatant was filtered through 0.22 µm PTFE filters into glass vials. Methanol extractions were repeated twice and extracts were pooled. Extracts were dried under vacuum at ambient temperature. Final extraction weight was determined, vials were sealed and stored at -20°C until analysis. Each biological replicate included an extraction |
Combined analysis:
| Analysis ID | AN003198 | AN003199 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 |
| Column | Phenomenex Kinitex C18 (50 x 2.1 mm,1.7um) | Phenomenex Kinitex C18 (50 x 2.1 mm,1.7um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo LTQ XL | Thermo LTQ XL |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | Area | Area |
Chromatography:
| Chromatography ID: | CH002365 |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | Phenomenex Kinitex C18 (50 x 2.1 mm,1.7um) |
| Flow Rate: | 0.35 ml/min |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS002976 |
| Analysis ID: | AN003198 |
| Instrument Name: | Thermo LTQ XL |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | voltage of 35 V m/z from 100 – 2000.00 MS1 data was processed using MZMine2 |
| Ion Mode: | POSITIVE |
| MS ID: | MS002977 |
| Analysis ID: | AN003199 |
| Instrument Name: | Thermo LTQ XL |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | voltage of -35 V m/z from 100 – 2000.00 MS1 data was processed using MZMine2 |
| Ion Mode: | NEGATIVE |
