 1.
[Figure ID: F3]
Absolute quantities for allergens and anti-nutritional proteins. The absolute quantities for the eight proteins of interest are presented. Data are grouped by cultivar with each separate bar representing the quantity of the protein for the indicated environment (location). Error bars are standard deviation between extraction replicates.
[Figure ID: F4]
Allergen totals by location. Values presented are the sum totals for all eight proteins measured. Data are grouped by location, and error bars are summed standard deviations for each allergen. Groups were ordered according to their average allergen sums.
[Figure ID: F5]
Environment versus cultivar biplot. Results of principal component analysis were visualized using a biplot. The x axis represents the direction of maximum variation through the data, which is called the first principle component direction. The y axis represents the direction of the next highest variation through the data, which is orthogonal to the first principle component and called the second principle component direction. Text in the body of the plot represents the samples. The name for each has two parts: location abbreviation followed by cultivar number. Correlations between allergens are represented by vectors terminating with text indicating the allergen they represent. Angles between these vectors, acute or obtuse, represent positive or negative correlation respectively.
[Figure ID: FA1]
Matrix dilution series. The linearity of response of endogenous peptides was assessed with constant AQUA™. A twofold dilution series from 2 to 0.125 μg/injection of soybean (Williams 82) digest was analyzed. The fractional changes, from undiluted through the twofold dilution series, both measured (via response ratio) and actual, were plotted to assess the slope and correlation values. Values on the x and y axes are the fraction of undiluted sample (2 μg/injection) for each successive dilution in the series (1, 0.5, 0.25, and 0.125 μg/injection). Technical variation for the experiment was on average, 8.86% when considering coefficients of variance for triplicate injections of all peptides at all dilutions.
ANOVA analysis of allergen levels.
|Beta conglycinin alpha||Site||5||15||15.38||<0.0001|
|Beta conglycinin alpha||Cultivar||3||15||0.92||0.4558|
|Gly m Bd 28k||Site||5||15||19.07||<0.0001|
|Gly m Bd 28k||Cultivar||3||15||4.47||0.0197|
For a given variable, genotypic variation, and environmental variation were quantified relative to residual variation using Analysis of Variance (ANOVA)-based F-test.
Allergen peptide analyte information.
|ID no.||Peptide name||Protein represented||Peptide sequence||Calculated (Da)||Observed (Th)||Glyma no.||Protein MW (g/mol)monoisotopic (M+H)1+|
|Gly m 6 (GLYCININ)|
|SA2||GlyG1-2||G1 subunit (Bx)||VLIVPQNFVVAAR||1425.857604||713.4||Glyma03g32030.1||55671.6|
|SA4||GlyG2-2||G2 subunit (A2)||NLQGENEEEDSGAIVTVK||1931.919212||966.5||Glyma03g32020.1, Glyma03g32020.2||54356.887636, 41045.114403|
|SA6||GlyG3-2||G3 subunit (A)||FYLAGNQEQEFLQYQPQK||2231.076708||1116.0||Glyma19g34780.1||54207.863911|
|SA7||GlyG4||G4 subunit (A5)||VESEGGLIQTWNSQHPELK||2152.066872||718.0||Glyma10g04280.1||63758.458975|
|Gly m 5 (BETA CONGLYCININ)|
|SA10||Bcon α||Alpha subunit||LITLAIPVNKPGR||1391.873253||464.6||Glyma20g28650.1, Glyma20g28650.2, Glyma20g28660.1||70263.437646, 63248.763394, 70263.437646|
|Gly mTI (KUNITZ TRYPSIN INHIBITOR)|
|SA11||KTI 3-1||KTI 3||FIAEGHPLSLK||1211.678251||404.6||Glyma08g45530.1||14517.404983|
|SA13||KTI 1||KTI 1||DTVDGWFNIER||1351.627676||676.3||Glyma01g10900.1||22432.404593|
|Gly m BD (P34)|
Soy peptide data – Peptide sequences, masses, and observed masses for all peptides used in absolute quantification experiments are presented. In addition, the name of the annotated Swiss-Prot entry (“Protein represented” column) that corresponds to the soybean genome sequence (“Glyma no.”, Schmutz et al., 2010; www.phytozome.org) used to design the peptide, is presented. In the case where multiple splice variants were present, the underlined information was used. The unch