|Detail of the metabolite profile
||Polar gas chromatography mass spectroscopy (GC-MS)
|Description for analysis method
||A portion of 1 mL aqueous extract was taken out and dried with speed vacuum for polar metabolite analysis. Dried polar extracts were methoximated in pyridine with 40 uL of 15.0 mg/mL methoxyamine?€?HCl, vetoxed thoughly and sonicated for 15 min at room temperature, followed by incubation at 50 ??C until the residue was resuspended (around 2 h). Metabolites were then derivatized with 30 uL of MSTFA+1% TMCS for 1 h at 50 ??C. The sample was subsequently transferred to a 200 uL glass insert and analysed by analysed using an Agilent 6890 GC coupled to a 5973 MSD scanning from m/z 50?€?650. Samples were injected at a 1:15 split ratio, and the inlet and transfer line were held at 280 ??C. Separation was achieved with a temperature programme of 80 ??C for 2 min, then ramped at 5 ??C/min to 315 ??C and held for 12 min, a 60 m DB-5MS column (J&W Scientific, 0.25 mm ID, 0.25 lm film thickness) and a constant flow of 1.0 ml/min.
|Species / Cultivar / Tissue
||Medicago sativa / leaf hopper susceptible line, Forage Genetics International / Trichome
|Description for biological material
||Trichomes were collected from 80 g F.W. stems of alfalfa plants and pooled together as one replicate for both metabolome and transcriptome experiments. After the first replicate samples were collected, the plants were cut back and grow for 2 weeks. Trichomes collected from these plants were the second replicate samples. And the 3rd replicates samples were collected from the cutting back of the 2nd replicate plants. The semi-quantification will be based on the amount of stems used to collect trichome. After shearing off trichomes, stems were simultaneously collected as a control for metabolite profiling.
||Don Sik Yang, Lloyd W. Sumner. Metabolic Profiling of trichomes from Potato Leafhopper susceptible and resistant alfalfa lines. (unpublished)
Plant Biology Division
The Samuel Roberts NOBLE Foundation, Inc.
Ardmore, OKLAHOMA, U.S.A - 73401.