Fig. S1a-i: MS 2 spectra of infection markers The putative identity of infection markers obtained by metabolite fingerprinting was confirmed by MS 2 fragmentation experiment by LC 1290 Infinity coupled with a 6540 UHD Accurate-Mass Q-TOF LC MS instrument with Agilent Jet Stream Technology as ESI source. m/z 359.1474 4 x10 0 0.2 0.4 0.6 0.8 1 1.2 1.4 1.6 1.8 2 2.2 2.4 2.6 -ESI Product Ion (2.799 min) Frag=160.0V [email protected] (567.2078[z=1] -> **) KF_Steffi_MSMSI_CE10_pol 419.1327 521.1997 329.1385 223.0604 133.0145 385.1472 Counts vs. Mass-to-Charge (m/z) 120 140 160 180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480 500 520 540 560 Lariciresinol glucoside (M+FA-H + ) - [M-H + ] - [M-H + +FA] - [M-Glc-H + ] - [M-Glc-CH 2 O-H + ] - [M-Glc-C 9 H 12 O- H + ] - 359.1474 - 162.0528, C 6 H 10 O 5 - 136.088, C 9 H 12 O - 30.0196, CH 2 O - 164.0825, C 10 H 12 O 2 195.0649 m/z 195.065 [C 10 H 11 O 4 ] - pporting Information Figure S1a O O-Glc O-C H 3 O H O-C H 3 O H
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Fig. S1a-i: MS 2 spectra of infection markers The putative identity of infection markers obtained by metabolite fingerprinting was confirmed by MS 2 fragmentation.
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Fig. S1a-i: MS2 spectra of infection markersThe putative identity of infection markers obtained by metabolite fingerprinting was confirmed by MS2 fragmentation experiment by LC 1290 Infinity coupled with a 6540 UHD Accurate-Mass Q-TOF LC MS instrument with Agilent Jet Stream Technology as ESI source.
Fig. S2: UPLC-TOF-MS based metabolite fingerprinting of V. longisporum infected Arabidopsis leaves.Arabidopsis plants of different time points after inoculation with V. longisporum (10, 21 and 35 dpi) and the corresponding control plants were harvested and the aerial parts were extracted by two phase partitioning. The fingerprint of metabolites of the polar extraction phase was generated by UPLC-TOF-MS analysis. A subset of high-quality 862 metabolite marker candidates (FDR <10-4) derived from the positive as well as the negative ionization mode were used for clustering and visualization by means of one-dimensional self-organizing map (1D-SOM). (a) 1D-SOM heat map after metabolite-based clustering with 20 prototypes. Horizontal and vertical dimensions correspond to prototypes and experimental conditions, respectively. The heat map colors represent average intensity values according to the color map on the right hand side. The width of each prototype column is proportional to the number of marker candidates assigned to this prototype. (b) Box plots of prominent markers from different clusters (cl.) which accumulate in V. longisporum inoculated plants (VL). Four samples per treatment from two independent experiments were analyzed. Boxes indicate the 25th (q1) and 75th (q3) intensity percentiles and median of the data. The whiskers indicate the range between q1-1.5(q3-q1) and q3+1.5(q3-q1), respectively.
control VL43
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Supporting Information Figure S3
Fig. S3: Lignin accumulation in V. longisporum infected vascular tissue of Arabidopsis.Cross-sections of hypocotyls and petioles of V. longisporum inoculated (VL) and control plants at 21 dpi. Material was paraffin embedded, hand sectioned and stained with phloroglucin/HCl for lignified tissue. Scale bar: 200 µm.
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Supporting Information Figure S4
Fig. S4: Infection of the C4H:F5H mutant with V. longisporum.Col 0 and C4H:F5H plants were infected by root dip infection and harvested at 21 dpi. (a) Pictures of infected and control plants at 21 dpi. (b) The relative leaf area was determined from pictures taken of the rosettes of the plants: The leaf area of infected plants was divided by the leaf area of control plants and the results of the mutant plants were set in relation to the value of the wild type. The data represent the mean values of five independent experiments ±SD. (c) Determination of VL-DNA in infected leaves. The mean values of ten samples of three independent experiments are shown ±SE.
Supporting Information Figure S5
Fig. S5: Sinapoyl glucose, coniferin and syringin in glycosyltransferase overexpressor plants and the fah1-2 mutant. Quantification of sinapoyl glucose and monolignols by HPLC-DAD of V. longisporum-inoculated (black bars) and control (white bars) plants. Syringin could not be quantified in the UGT72E2-OE because of high coniferin peaks. Each data point represents the mean value of eight samples from two independent experiments + SD. (*, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001 )
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Fig S6: Influence of ferulic and sinapic acid on V. longisporum growth.PDB plates were supplemented with ferulic acid and sinapic acid in the indicated concentrations. After inoculation the fungus was grown for 18 days on the plates. Pictures of colonies were taken under a binocular and the colony area was determined. The data represent the mean values ±SD of seven different plates of two independent experiments. Asterisks indicate significant differences between control and supplemented plates according to student’s t-test (*, P ≤ 0.05; ***, P ≤ 0.001).
Supporting Information Figure S6
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Supporting Information Table S1
Table S1: Primers used for the quantitative RT-PCR.
QuantPrime: Samuel Arvidsson, Miroslaw Kwasniewski, Diego M. Riano-Pachon and Bernd Mueller-Roeber: QuantPrime - a flexible tool for reliable high-throughput primer design for quantitative PCR. BMC Bioinformatics 2008, 9:465
name ATG Primer referencePhenylalanine ammonia lyase 1 (PAL1)
Table S2: Data matrix of 826 high quality marker candidates (false discovery rate<10 -4) obtained by metabolite fingerprinting (UPLC-ESI-TOF-MS analysis) of mock treated (control) or V. longisporum infected (VL) leaves of Arabidopsis (10, 21, 35 dpi) (see separate file).
Supporting Information Table S3
Table S3: Infection markers identified by metabolite fingerprinting (UPLC-ESI-TOF-MS analysis) and verified by UHPLC-ESI-QTOF-MS2 analysis or co-elution (see separate file).
Linie Atg plant line literature effected gene susceptibility
fah1-2 At4g36220 CS6172 (Chapple et al. 1992) ferulate-5-hydroxylase defect