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Figure S1. The %5H- and %benzodioxane-units in lignins of different mutant and transgenic plant lines, reported in literature and in this study. Data from previously reported NMR studies was re-evaluated here (from uncorrected volume integrals). %5H is 5H/(S+G+5H) and %benzodioxane levels is J/(A+B+C+J) – the relatively minor dibenzodioxocins D were neglected here (see Fig. 3 for structures of G, 5H and S and Figure S3 for structures of A, B, C, D and J). Note that these spectra were run over many years on different instruments and under different acquisition conditions. The values are nevertheless reasonably comparable and, interestingly, there is a fairly regular (almost linear) correlation between %benzodioxanes and %5H units (See graph). Thioacidolysis releases 5H monomers from only a fraction of the benzodioxane units. 1In the paper of Marita et al. (2003) 38% benzodioxane inter-unit linkages were noted as structures (d) were also included in the calculation; 2The COMT-silenced poplars are the same lines; 3The C4H:F5H1 Arabidopsis plants are the same lines; n.r. not reported.

Figure S2. Partial short-range 13C–1H (HSQC) spectra (aromatic regions) of acetylated enzyme lignins isolated from agar-grown plants (a–e) and soil-grown plants (f–k). The S-unit integral estimate comes from subtracting the 5H2 contour integral from the S2/6+5H6 integral; the G-unit integral is determined directly from the resolved G2 component, as in the WT. For those genotypes that were grown both in soil and on agar, no major differences in lignin composition are seen between the two growth conditions.

Figure S3. Partial short-range 13C–1H (HSQC) spectra (aliphatic regions) of acetylated enzyme lignins isolated from agar-grown plants (a–e) and soil-grown plants (f–k), with color-coded assignments for units A–D and the benzodioxane units J resulting from 5-hydroxyconiferyl alcohol incorporation. For those genotypes that were grown both in soil and on agar, no major differences in lignin composition are seen between the two growth conditions.

Figure S4. UV-Vis absorption chromatogram of inflorescence stem extracts of comt C4H:F5H1 and WT plants. comt C4H:F5H1 plants accumulate large amounts of 5-hydroxylated phenylpropanoids (1–38), the identities of which are given in Table 3. Several peaks that are highly abundant in WT are given, cis- and trans-sinapoyl malate (SM), sinapoyl glucose (SG) and flavonol glycosides (FG1, 3-O-Rha(1->2)Glu-7-O-Rha-kaemferol; FG2, 3-O-Glu-7-O-Rha-kaemferol; FG3, 3,7-di-O-Rha-kaemferol; FG4, kaemferol rhamnoside; FG5, 3-O-Rha(1->2)Glu-7-O-Rha-quercitin; FG6, 3-O-Glu- 7-O-Rha-quercitin). Grey, profile of WT; red, profile of comt C4H:F5H1.

Figure S5. MS spectra of thirty eight 5-hydroxylated phenylpropanoids that were identified in comt C4H:F5H1 plants, with the reasoning for structural identification.

Table S1. Primers used in qRT-PCR experiments. 1For ACTIN (ACT) genes, the primers amplify ACT2 (At3g18780) and ACT8 (At1g49240) simultaneously (Charrier et al., 2002).

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