SE125:/MS1

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Sample Set Information

ID TSE4
Title A novel method for single-grain-based metabolic profiling of Arabidopsis seed
Description In plant metabolomics, metabolite contents are often normalized by sample weight. However, accurate weighing of very small samples, such as individual Arabidopsis thaliana seeds (approximately 20 µg), is difficult, which may lead to irreproducible results.
Authors Yuji Sawada, Hirokazu Tsukaya, Yimeng Li, Muneo Sato, Kensuke Kawade, Masami Yokota Hirai
Reference Sawada et al. (2017) Metabolomics 13:75
Comment


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The raw data files are available at DROP Met web site in PRIMe database of RIKEN.

Analytical Method Details Information

ID MS1
Title LC-MS/MS (triple quadrupole)
Instrument Waters XEVO-TQS
Instrument Type
Ionization ESI
Ion Mode Positive
Description Two sample preparation methods were employed for comparison. For the single-grain-based analysis, a visually normal seed was selected from each of the 12 individual plants under a microscope. After the estimation of seed volume as described above, each seed was placed in a separate well of a 96-well plate using a Vacuseed (Polaris Instruments Ltd, Cambridge, U.K.) with a 3-mm zirconia bead. The metabolites were extracted using 500 µL extraction solvent (80% methanol, 0.1% formic acid, 16.8 nmol/L lidocain, and 105 nmol/L 10-camphorsulfonic acid as internal standards) with a Multi-beads Shocker (Shake Master NEO, Bio Medical Science) at 1000 rpm for 10 min. After centrifugation, 250 µL of the extract was transferred to a new plate, dried, dissolved in 250 µL of ultra-pure water, and filtered using Whatman® UNIFILTER® plates 384 (GE Healthcare). The sample preparation process was automatically performed by a liquid handling system (Microlab Star Plus, Hamilton) [see widely targeted metabolomics section in PRIMe (http://prime.psc.riken.jp/) (Sakurai et al. 2013)]. 1 µL of the solution extract (final concentration = ca. 40 µg/mL) prepared from a seed (ca. 20 µg in the case of diploid) was subjected to widely targeted metabolomics using LC-QqQ-MS (UPLC-TQS, Waters).


For the weight-based analysis, approximately 200 seeds, weighing 4 ± 0.4 mg, from each of 12 plants, were collected using a Seed Spoon (BMS-SS200, Bio Medical Science). Seeds were accurately weighed, placed in a 2-mL tube with a 5-mm zirconia bead and proportional volume of the extraction solvent (4.0 mg/mL), and crushed using a Multi-beads Shocker at 1000 rpm for 10 min. After centrifugation, the extracts were diluted to 40 µg/mL with the extraction solvent. A 250 µL volume of the extract was transferred to a 96-well plate, dried, dissolved in 250 µL of ultra-pure water, and filtered using Whatman® UNIFILTER® plates 384. 1 µL of solution extract (40 µg/mL) was injected into an LC-QqQ-MS (UPLC-TQS, Waters) (Fig. 2).

Comment_of_details All raw data (Waters LC-MS/MS data) and metadata are downloadable from DROP Met (http://prime.psc.riken.jp/) in PRIMe(Sakurai et al. 2013).


Link icon database.png Link icon dropmet.png

The raw data files are available at DROP Met web site in PRIMe database of RIKEN.

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