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Sample Set Information

ID TSE1307
Title Induced accumulation of glucuronosyldiacylglycerol in tomato and soybean under phosphorus deprivation.
Description Glucuronosyldiacylglycerol (GlcADG) is a plant glycolipid that accumulates in Arabidopsis and rice in response to phosphorus (P) starvation. It has been suggested that GlcADG functions to mitigate the stress induced by P depletion. Biosynthesis of GlcADG requires sulfolipid (SQDG) synthase, which is coded for in plant genomes. This indicates the possibility that GlcADG may be a general constituent of membrane lipids in plants. In this study, we investigated the SQDG synthases found in the genomes of higher plants, ferns, mosses, algae and cyanobacteria. In addition, we analyzed GlcADG accumulation, and the expression of SQDG synthase homologs in tomato and soybean plants grown under P-limited conditions. LC-MS analysis of lipids from these plants confirmed that GlcADG accumulated during P deprivation, as previously observed in Arabidopsis and rice. We also observed upregulation of SQDG synthase transcripts in these plants during P deprivation. These data suggest that GlcADG is present not only in model plants, but also in various other plant species, and that this lipid molecule performs an important physiological function as a mitigator of P-deprivation stress in plants.
Authors Okazaki Y, Nishizawa T, Takano K, Ohnishi M, Mimura T, Saito K.
Reference Physiol Plant. 2015 Sep;155(1):33-42. doi: 10.1111/ppl.12334. Epub 2015 Mar 12.

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Sample Information

Title Soybean
Organism - Scientific Name Glycine max
Organism - ID NCBI taxonomy:3847
Compound - ID
Compound - Source
Preparation Soybean seeds (Glycine max (L.) Merr. cv. Enrei) were agitated in 2% PPM solution, and sown in wet potting mix consisting of 2:1 (v/v) PRO-MIX BX (Premier Tech Horticulture, Rivière-du-Loup (Québec), Canada) and vermiculite (VS kakou, Minato-ku, Tokyo, Japan), and grown under a 28°C/20°C day/night temperature cycle with a 12 h photoperiod. The roots of 10-day-old soybean plants were gently washed to remove the soil, then transferred to rockwool, where the P levels were controlled as described above.
Sample Preparation Details ID

Analytical Method Information

Method Details ID MS1
Sample Amount

Analytical Method Details Information

Title Lipidomic analysis
Instrument LC, Waters Acquity UPLC system; MS, Waters Xevo G2 Q-Tof
Instrument Type UPLC-QTOF-MS
Ionization ESI
Ion Mode positive
Description Total lipids were extracted using a mixture of methyl tert-butyl ether (MTBE) and methanol, as previously reported (Matyash et al. 2008, Giavalisco et al. 2011), with slight modifications. The frozen plant samples were milled to a fine powder under cryogenic conditions, as described previously (Okazaki et al. 2013a).

Total lipids were extracted by adding 16 volumes of extraction solvent (3:1, MTBE/methanol [v/v] with 1 μM 1,2-didecanoyl-sn-glycero-3-phosphocholine [Sigma-Aldrich] added as an internal standard) to the powdered plant sample while in liquid nitrogen, followed by vigorous vortexing. To this mixture, four volumes of water was added, followed by vigorous mixing on a sample tube mixer for 5 min at room temperature, incubation on ice for 15 min, and centrifugation at 1000 g at 5°C for 5 min. After centrifugation, a 100-μl aliquot of the supernatant was collected and evaporated to dryness in a centrifugal concentrator. The residue was reconstituted in 125 μl of ethanol and centrifuged at 10 000 g at 5°C for 15min. The supernatant was used immediately for lipid analysis.
Lipid analysis was conducted using liquid chromatography quadrupole time-of-flight mass spectrometry on a Waters Xevo G2 Q-TOF MS combined with a Waters ACQUITY UPLC system, as reported previously (Kimbara et al. 2013). The conditions for recording MS/MS in positive-ion mode were as follows: scan range, m/z 100–1200; capillary, 3.0 kV; sampling cone, 20; extraction cone, 4.0; source temperature, 120°C; desolvation temperature, 450°C; cone gas flow, 50 l h−1; desolvation gas flow, 600 l h−1; collision energy ramp start, 20 eV; collision energy ramp end, 50 eV. The conditions for recording MS/MS in negative-ion mode were as follows: scan range, m/z 100–1200; capillary, 2.5 kV; sampling scone, 40; extraction cone, 4.0; source temperature, 120°C; desolvation temperature, 450°C; cone gas flow, 50 l h−1; desolvation gas flow, 600 l h−1; collision energy ramp start, 20 eV; and collision energy ramp end, 50 eV.

Detection of GlcADG in tomato and soybean
Ten-day-old tomato and soybean seedlings were transplanted to rockwool, and nutrients were supplied by the application of liquid nutrient medium. For the P-limitation treatment, liquid medium without potassium phosphate was used. After a 14-day incubation period, lipids were extracted from the leaves and analyzed by LC-Q-TOF-MS.

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