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. 2022 Oct 12;13:1000819. doi: 10.3389/fpls.2022.1000819

Figure 2.

Figure 2

Detailed molecular characterization of camphor-0 M3 lines. Levels of artemisinin-related and other selected mono- and sesquiterpenes (A) were quantified by gas chromatography–mass spectrometry (GC-MS) (i), (iii), (viii) - (xii) and ultra-high performance liquid chromatography–mass spectrometry (UPLC-MS) (ii), (iv)-(vii) analysis of extracts from fresh leaves L1-5 (juvenile), L7-9 (expanding), L11-13 (mature) as counted from the apical meristem from 12-weeks old glasshouse-grown Artemis F1 (blue bars) and selected camphor-0 M3 (orange bars); error bars – SE (n=5). nd - not detectable. GC-MS data for camphor were converted from internal standard (IS) and dry weight (DW) normalised peak areas to µg/mg extracted dry weight (vii) against a standard curve run with pure camphor. For both (A, B) letters represent Tukey’s range test results after one way ANOVA or REML. Groups not sharing letters indicate statistically significant differences. (B) Level of total volatiles measured by GC-MS in three types of leaf material as indicated in (A). (C) Level of FPP measured in extracts from fresh juvenile leaves L1-5 Artemis F1 (blue bars) and selected camphor-0 M3 (orange bars); FW - fresh weight, error bars – SE (n=3), *- statistically significant difference (t-test) between Artemis and camphor-0 at p<0.05. (D) Summary of metabolite changes in the camphor-0 glandular secretory trichomes (GSTs), full arrows – known enzymatic steps, dotted arrows – potential non enzymatic conversions, full green arrows – metabolite changes (all leaf stages). Metabolite abbreviations: GPP - geranyl diphosphate, FPP – farnesyl diphosphate, A-4,11-D – amorpha-4,11-diene, DHAA - dihydroartemsinic acid, DHAAOOH- dihydroartemisinic acid tertiary hydroperoxide, DHEDB – dihydro-epi-deoxyarteanniun B, Enzyme abbreviations: IDI - Isopentenyl Diphosphate Isomerase, GPS - Geranyl diPhosphate synthase, BPS - Bornyl diPhosphate synthase, FPS- Farnesyl diPhosphate Synthase. Artemisinin pathway: AMS – amorpha-4,11-diene synthase, CYP71AV1 - amorpha-4,11-diene C-12 oxidase, CPR – cytochrome P450 reductase, DBR2 - artemisinic aldehyde Δ 11 (13) reductase, ALDH1 - aldehyde dehydrogenase. Question marks indicate putative active transport systems operating in A annua GSTs.