Return to study ST002330 main page
MB Sample ID: SA229675
| Local Sample ID: | HILIC-RN7-neg |
| Subject ID: | SU002417 |
| Subject Type: | Plant |
| Subject Species: | Brassica napus |
| Taxonomy ID: | 3708 |
Select appropriate tab below to view additional metadata details:
Sample Preparation:
| Sampleprep ID: | SP002423 |
| Sampleprep Summary: | Approximately 100 mg of fresh weight of root tissue samples (n=3) were aliquoted from finely homogenized frozen root tissues, placed into 2 mL safe-lock centrifuge tubes with two zirconium oxide beads, flash frozen in liquid nitrogen and stored at − 80 °C until further processing. To accommodate for different extraction procedures, two sample sets were prepared from the same bulk to independently analyze the content of primary metabolites (free amino acids, sugars and sugar phosphates, organic acids), and secondary metabolites (glucosinolates) using High Performance Liquid Chromatography – High-Resolution Accurate Mass – Full Scan Mass Spectrometry (HPLC-(HRAM)-MS). Samples for extraction of primary and secondary metabolites were spiked with 10ng of two labeled aromatic CKs (13C5-oT and 2H7-BAR) and extracted with ice-cold methanol (methanol-water [8:2, v/v]) following procedures of Chen et al 3 with modifications. Each filtered extract was split in half and each 500 µL transferred to a new 2 mL tube. Divided extracts were evaporated to dryness at ambient temperature in a speed vac concentrator. Sample residues designed for the analysis of primary metabolites were redissolved in 500 µl of 90% acetonitrile (acetonitrile: water, v/v) and samples for the analysis of glucosinolates were redissolved in 500 µL of 5% acetonitrile with 0.1% formic acid (acetonitrile: formic acid: water, v/v/v). Additionally, sample mixtures composed of 10uL of each sample extract were prepared separately for primary and secondary metabolite analysis and used to generate MS/MS data for compound identification. All samples were filtered using 0.2 µm PVDF spin filter with 2 mL receiver tubes (InnoSep Spin, Canadian Life Sciences, Peterborough, Canada) and transferred to insert-equipped 2 mL HPLC vials. A volume of 25 μL of each sample was injected into a Dionex UltiMate 3000 HPLC coupled to a QExactive Orbitrap mass spectrometer. |
| Sampleprep Protocol Filename: | Protocol_methods-canola |
| Extraction Method: | 80% MeOH |
| Extract Cleanup: | 0.2 µm PVDF spin filter with 2 mL receiver tubes |
| Extract Storage: | -20℃ |
| Sample Resuspension: | Primary metabolites - 90% ACN, glucosinolates - 5% ACN |
| Sample Spiking: | two labeled aromatic CKs (13C5-oT and 2H7-BAR) |
