SE193:/MS1

From Metabolonote
jump-to-nav Jump to: navigation, search

Sample Set Information

ID TSE1352
Title Metabolic Reprogramming in Leaf Lettuce Grown Under Different Light Quality and Intensity Conditions Using Narrow-Band LEDs
Description Light-emitting diodes (LEDs) are an artificial light source used in closed-type plant factories and provide a promising solution for a year-round supply of green leafy vegetables, such as lettuce (Lactuca sativa L.). Obtaining high-quality seedlings using controlled irradiation from LEDs is critical, as the seedling health affects the growth and yield of leaf lettuce after transplantation. Because key molecular pathways underlying plant responses to a specific light quality and intensity remain poorly characterised, we used a multi-omics–based approach to evaluate the metabolic and transcriptional reprogramming of leaf lettuce seedlings grown under narrow-band LED lighting. Four types of monochromatic LEDs (one blue, two green and one red) and white fluorescent light (control) were used at low and high intensities (100 and 300 μmol·m−2·s−1, respectively). Multi-platform mass spectrometry-based metabolomics and RNA-Seq were used to determine changes in the metabolome and transcriptome of lettuce plants in response to different light qualities and intensities. Metabolic pathway analysis revealed distinct regulatory mechanisms involved in flavonoid and phenylpropanoid biosynthetic pathways under blue and green wavelengths. Taken together, these data suggest that the energy transmitted by green light is effective in creating a balance between biomass production and the production of secondary metabolites involved in plant defence.
Authors Kazuyoshi Kitazaki, Atsushi Fukushima, Ryo Nakabayashi, Yozo Okazaki, Makoto Kobayashi, Tetsuya Mori, Tomoko Nishizawa, Sebastian Reyes-Chin-Wo, Richard W. Michelmore, Kazuki Saito, Kazuhiro Shoji & Miyako Kusano
Reference Scientific Reports, volume 8, Article number: 7914 (2018)
Comment


Link icon article.png

Analytical Method Details Information

ID MS1
Title GC-TOF-MS
Instrument GC Agilent 6890N gas chromatograph / MS Pegasus IV TOF mass spectrometer
Instrument Type
Ionization EI
Ion Mode Positive
Description Extraction and derivatization for GC-TOF-MS

Each frozen sample with a 5-mm zirconia bead was extracted with 400 fold amount of solvent (methanol/chloroform/water [3:1:1 v/v/v]) containing 10 stable isotope reference compounds at 4°C in a mixer mill (MM301; Retsch, Haan, Germany) at a frequency of 15 Hz. Each isotope compound was adjusted to a final concentration of 15 ng per 1-µl injection volume. After 5-min centrifugation at 15,100 × g, a 200-µl aliquot of the supernatant was transferred to a glass insert vial. The extracts were evaporated to dryness in an SPD2010 SpeedVac® concentrator (Thermo Fisher, Scientific, Waltham, MA, USA). We used extracts from 0.5-mg DW samples for derivatization, i.e., methoxymation and silylation. For methoxymation, 30 µl of methoxyamine hydrochloride (20 mg/ml in pyridine) were added to the sample. After 22.5 h of derivatization at room temperature the sample was trimethylsilylated for 1 h using 30 µl of MSTFA at 37°C with shaking. All derivatization steps were performed in a vacuum glove box VSC-1000 (Sanplatec, Osaka, Japan) filled with 99.9995% (G3 grade) dry nitrogen.

GC-TOF-MS conditions
Using the splitless mode of a CTC CombiPALautosampler (CTC Analytics, Zwingen, Switzerland), 1 µl of each sample (equivalent to 5.6 µg DW) was injected into an Agilent 6890N gas chromatograph (Agilent Technologies, Wilmingston, DE, USA) featuring a 30 m × 0.25 mm inner diameter fused-silica capillary column and a chemically bound 0.25-µl film Rxi-5 Sil MS stationary phase (RESTEK, Bellefonte, PA, USA) with a tandem connection to a fused silica tube (1 m, 0.15 mm). An MS column change interface (msNoVent-J; SGE, Yokohama, Japan) was used to prevent air and water from entering the MS during column change-over. Helium was the carrier gas at a constant flow rate of 1 ml min-1. The temperature program for GC-MS analysis started with a 2-min isothermal step at 80°C followed by 30°C temperature-ramping to a final temperature of 320°C that was maintained for 3.5 min. The transfer line and the ion source temperatures were 250 and 200°C, respectively. Ions were generated by a 70-eV electron beam at an ionization current of 2.0 mA. The acceleration voltage was turned on after a solvent delay of 222 sec. Data acquisition was on a Pegasus IV TOF mass spectrometer (LECO, St. Joseph, MI, USA); the acquisition rate was 30 spectras-1 in the mass range of a mass-to-charge ratio of m/z = 60–800. Supplementary Material 4 Alkane standard mixtures (C8 - C20 and C21 - C40) purchased from Sigma-Aldrich (Tokyo, Japan) were used for calculating the retention index (RI) 1 . For quality control we injected methylstearate into every 6th sample. The sample run order was randomized in single-sequence analyses. We analyzed the standard compound mixtures using the same sequence analysis procedures.

Comment_of_details


Personal tools
View and Edit Metadata
Variants
Views
Actions