Supplementary Table I. Comparison of the sugar linkage groups in the pectins of isolated cell walls extracted with 0.5% ammonium oxalate. Three control, C1, C2 and C3 and three transgenic TL0, TL1 and TL2 samples were used.
Sugar C1C2C3TL0TL1TL2
and Linkage
mole%*
Fucose:
t-Fuc0.40.30.40.80.60.7
Rhamnose:
t-Rha0.20.30.30.30.20.3
2-Rha2.62.22.13.12.73.2
2,3-Rha0.20.20.20.20.10.2
2,4-Rha0.20.91.21.21.21.3
Arabinose:
t-Araf1.11.91.41.40.91.4
t-Arap0.2tr0.2trtrtr
2-Araf0.10.50.10.10.10.1
3-Araf0.30.80.50.60.50.5
4-Arap0.40.30.20.40.40.5
5-Araf3.63.63.55.04.33.2
2,5-Araf1.52.31.62.11.91.5
3,5-Araf0.60.60.60.80.70.5
Supplementary Table I, continued ______
Sugar C1C2C3TL0TL1TL2
and Linkage
______
mole%
Xylose:
t-Xyl2.41.62.42.31.32.4
2-Xyl1.30.23.62.61.41.5
4-Xyl7.82.47.85.16.95.3
2,4-Xyl6.63.26.710.56.26.4
3,4-Xyl0.40.22.30.20.30.2
Mannose:
t-Man0.20.1tr0.30.30.2
4-Man2.31.53.73.33.03.5
4,6-Man0.40.20.40.40.70.5
Galactose:
t-Gal2.20.92.70.72.22.0
2-Gal0.20.10.4tr0.20.2
3-Gal0.70.20.70.10.60.5
4-Gal1.41.90.80.30.90.7
6-Gal1.40.52.00.31.51.3
2,4-Gal0.60.51.20.10.71.0
3,4-Gal0.60.30.60.10.50.4
3,6-Gal1.10.32.00.41.21.1
Supplementary Table I, continued ______
Sugar C1C2C3TL0TL1TL2
and Linkage
______
mole%
Galacturonic Acid:
t-GalA1.41.11.41.51.81.7
4-GalA36.439.427.929.733.636.6
2,4-GalA5.64.03.94.15.23.6
3,4-GalA8.57.66.47.910.19.7
Glucose:
t-Glc0.41.11.41.51.81.7
4-Glc2.514.75.63.33.83.2
3,4-Glc0.31.40.80.30.40.3
4,6-Glc0.31.20.60.40.71.4
Glucuronic Acid:
t-GlcA3.12.22.83.82.83.0
*Pectinaceous materials extracted from isolated cell walls by hot ammonium oxalate were filtered through glass fiber filter mats, dialyzed extensively against deionized water, and then the uronosyl residues were carboxyl-reduced with CMC and NaBD4 as described by Carpita and McCann (1996). After dialysis, the materials were freeze-dried, and alditol acetates and partly methylated alditol acetates were prepared, and the derivatives were separated by gas-liquid chromatography, as described by Gibeaut and Carpita (1991). The sugar linkage groups were identified by electron-impact mass spectrometry as described by Carpita and Shea (1989). Carboxyl-reduced uronosyl residues were distinguished from their respective neutral sugar as 6,6-dideutero-hexitol acetates, and mole% was determined, with accounting for 13C-spillover, as described by Kim and Carpita (1992). Values are the means of two samples; variance was typically within 5% of mean. The mole% of each monosaccharide and carboxyl-reduced uronic acid was determined by alditol acetates, using myo-inositol as an internal standard (not shown). tr = trace amounts less than 0.5%
Supplementary Table II. Comparison of sugar and linkage distribution of cross-linking glycans from depectinated cell walls extracted by 1 M NaOH. Three control, C1, C2 and C3 and three transgenic TL0, TL1 and TL2 samples were used.
SugarC1C2C3TL0TL1TL2
and Linkage
______
mole%*
Fucose:
t-Fuctrtrtrtrn.d.n.d.
Rhamnose:
t-Rhatrtr0.1trn.d.0.1
2-Rha0.40.70.40.30.30.3
2,4-Rha0.60.20.60.50.50.5
Arabinose:
t-Araf0.50.70.40.50.40.4
t-Arap0.60.40.60.50.81.0
2-Araftr0.1trtrtr0.2
3-Araf0.30.2trtrtrtr
5-Araf1.21.41.51.21.11.1
2,5-Araftr0.30.20.20.50.1
3,5-Araf0.50.30.40.50.30.3
Supplementary Table II, continued ______
Sugar C1C2C3TL0TL1TL2
and Linkage
______
mole%
Xylose:
t-Xyl0.60.70.50.81.00.8
4-Xyl85.077.084.183.383.783.5
2,4-Xyl8.18.27.69.69.69.7
3,4-Xyl0.70.80.71.20.70.8
Mannose:
t-Man0.10.10.10.10.10.1
4-Man0.20.20.1tr0.1tr
4,6-Mantrtrtrtrtrtr
Galactose:
t-Gal0.20.40.10.10.10.1
4-Gal0.10.20.10.20.10.1
6-Gal0.10.20.1tr0.10.1
3,4-Gal0.20.40.20.10.10.1
3,6-Gal0.10.20.10.1trtr
Glucose:
t-Glc0.10.40.20.10.10.2
4-Glc0.46.31.70.50.40.4
4,6-Glc0.20.70.20.10.10.1
*Cross-linking glycans extracted from depectinated cell walls by 1 M NaOH (containing 3 mg/ml NaBH4) were filtered through glass fiber filter mats, neutralized with gl. acetic acid, dialyzed extensively against deionized water, and freeze-dried. Alditol acetates and partly methylated alditol acetates were prepared, separated, and identified as described in Table 1. The mole% of each monosaccharide and uronic acid was determined by alditol acetates, using myo-inositol as an internal standard (not shown) and applied to the linkage distribution in methylation analysis. tr = trace amounts less than 0.05%; n.d. = not detected
Supplementary Table III. Comparison of sugar and linkage distribution of cross-linking glycans from depectinated and 1 M NaOH-treated cell walls extracted by 4 M NaOH. Three control, C1, C2 and C3 and three transgenic TL0, TL1 and TL2 samples were used.
______
SugarC1C2C3TL0TL1TL2
and Linkage
______
mole%*
Fucose:
t-Fuc0.10.20.10.10.10.1
Rhamnose:
t-Rha0.1tr0.10.10.10.1
2-Rha0.40.40.40.30.40.3
3-Rha0.3trtr0.30.40.3
2,4-Rha0.30.50.60.50.50.5
Arabinose:
t-Araf0.80.90.90.80.60.7
t-Arap0.30.20.30.20.20.2
2-Araf0.10.10.10.10.10.1
3-Araf0.10.40.20.20.20.2
4-Arap0.20.20.50.20.20.3
5-Araf1.01.21.11.81.71.4
2,5-Araf0.50.40.30.30.40.3
3,5-Araf0.40.20.40.20.30.6
Supplementary Table III, continued
______
SugarC1C2C3TL0TL1TL2
and Linkage
______
mole%
Xylose:
t-Xyl0.60.40.60.90.60.8
4-Xyl68.660.568.972.474.874.2
2,4-Xyl10.18.09.712.111.310.8
3,4-Xyl1.91.71.72.62.02.1
Mannose:
t-Man0.10.10.10.1tr0.1
4-Man4.84.64.91.40.91.3
4,6-Man0.30.40.30.10.10.1
Galactose:
t-Gal0.60.70.10.40.50.5
2-Gal0.50.40.10.20.20.2
4-Gal0.30.2tr0.30.20.2
6-Gal0.40.40.10.50.50.4
2,4-Gal0.30.30.10.1tr0.1
3,4-Gal0.30.40.10.20.40.3
3,6-Gal0.20.20.10.20.20.2
Supplementary Table III, continued
______
Sugar C1C2C3TL0TL1TL2
and Linkage
______
mole%
Glucose:
t-Glc0.20.70.30.10.10.1
4-Glc5.012.75.52.31.82.3
4,6-Glc0.81.80.70.60.70.6
*Cross-linking glycans extracted from depectinated cell walls by 4 M NaOH (containing 3 mg/ml NaBH4) were filtered through glass fiber filter mats, neutralized with gl. acetic acid, dialyzed extensively against deionized water, and freeze-dried. Alditol acetates and partly methylated alditol acetates were prepared, separated, and identified as described in Table 1. The mole% of each monosaccharide and uronic acid was determined by alditol acetates, using myo-inositol as an internal standard (not shown) and applied to the linkage distribution in methylation analysis. tr = trace amounts less than 0.5%.
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