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Analysis of barley shrunken grain mutants has identified lines with a novel high amylose starch phenotype. The causal mutation is located at the sex6 locus on chromosome 7H, suggesting the starch synthase IIa (ssIIa) gene as a candidate gene altered by the mutation. Consistent with this hypothesis, no evidence of SSIIa protein expression in either the starch granule or soluble fractions of the endosperm was found. Sequences of the starch synthase IIa gene, ssIIa, from three independent sex6 lines showed the presence of a stop codon preventing translation of the ssIIa transcript in each line. Perfect segregation of the starch phenotype with the presence of stop codons in the ssIIa gene was obtained, providing strong evidence for the lesion in the ssIIa gene being the causal mutation for the sex6 phenotype. The loss of SSIIa activity in barley leads to novel and informative phenotypes. First, a decrease in amylopectin synthesis to less than 20% of the wild-type levels indicates that SSIIa accounts for the majority of the amylopectin polymer elongation activity in barley. Secondly, in contrast to high amylose starches resulting from branching enzyme downregulation, the sex6 starches have a shortened amylopectin chain length distribution and a reduced gelatinisation temperature. Thirdly, the mutation leads to pleiotropic effects on other enzymes of the starch biosynthesis pathway, abolishing the binding of SSI, branching enzyme IIa and branching enzyme IIb to the starch granules of sex6 mutants, while not significantly altering their expression levels in the soluble fraction.

Cereal Chem. 74(1):63-71The starch of wheat (Triticum aestivum L.) flour affects food product quality due to the temperature-dependent interactions of starch with water during gelatinization, pasting, and gelation. The objective of this study was to determine the fundamental basis of variation in gelatinization, pasting, and gelation of prime starch derived from seven different wheat cultivars: Kanto 107, which is a partial waxy mutant line, and six near-isogenic lines (NILs) differing in hardness. Complete pasting curves with extended 16-min hold at 93°C were obtained using the Rapid Visco Analyser (RVA). Apparent amylose content ranged from 17.5 to 23.5%; total amylose content ranged from 22.8 to 28.2%. Starches exhibited significant variation in onset of gelatinization. However, none of the parameters measured consistently correlated with onset or other RVA curve parameters that preceded peak paste viscosity. Peak paste viscosity varied from 190 to 323 RVA units (RVU). Higher peak, greater breakdown, lower final viscosity, negative setback, and less total setback

Electrophoretic characterisation of fractions collected from gluten protein extracts subjected to size-exclusion high-performance liquid chromatographyThe electrophoretic analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE; reduced and unreduced) of fractions, collected from a size exclusion-high performance liquid chromatography (SE-HPLC) separation of gluten proteins using a column with pore size of around 400A, showed clear resolution for the seven elution ranges studied in two Australian bread wheat lines. Polymeric proteins -high molecular weight (HMW) glutenin subunits, low molecular weight (LMW) glutenin subunits, HMW albumins and some modified m-gliadins -appeared exclusively in the region within the first peak of the chromatogram (fractions 1 to 5), the limit being a region that resolved as a small peak before the large peak of gliadins and where some o-gliadins eluted. A larger proportion of HMW glutenin subunits and B subunits contributed to polymer formation of higher molecular weight.The polymer size decreased as the proportion of the other protein components increased. IntroductionWheat endosperm proteins have been extensively studied due to their viscoelastic properties that give wheat flours unique baking characteristics. They are usually classified according to their solubility, amino acid composition, size or polymerising behaviour. Wheat proteins may be separated into albumin (soluble in water), globulin (soluble in salt solution), gliadin (70% ethanol soluble) and glutenin (insoluble in ethanol) [l]. From a functional point of view, they are also grouped into polymeric protein (mainly glutenin) held together by interchain disulphide bonds, and monomeric protein (gliadins and albumins/globulins) [2], in which the only disulphide bonds are intrachain. Statistical relationships between these groups (particularly glutenins) and breadmaking quality have been established and, as a general result, close correlations have been found [2,3]. To reach these conclusions, wheat proteins have been characterised in detail using a wide range of methodologies, particularly over the last twenty-five years.Electrophoretic techniques using SDS-PAGE have been essential in determining the importance of glutenin polypeptides in wheat quality [4], but prior to electrophoresis in an SDS-PAGE gel, the glutenin polymer needs to be broken into subunits by the reduction of the disulphide bonds that build the polymeric structure. Four different types of subunits are found: high molecular weight (HMW) glutenin subunits, low molecular weight (LMW) glutenin subunits, HMW albumin subunits (mostly

A new method is described for determining the starch content of cereal products. It involves the solubilization of starch with a thermostable a-amylase followed by the complete hydrolysis with an amyloglucosidase. The glucose is then determined by the hexokinase/glucose-6 phosphate dehydrogenase procedure. By using an aamylase at high temperature, an autoclave is not required to gelatinize and solubilize the starch prior to amyloglucosidase treatment. a-Amylase is added to the sample before heating and degradation begins as soon as the gelatinization temperature is reached, before retrogradation can occur. The procedure was found to be superior to either treatment with acid or alkali, methods which also do not require an autoclave. Starkebestimrnung unter Verwendung thermostabiler aAmylasen. Eine neue Methode zur Bestimmung des Starkegehaltes von Getreideprodukten wird beschrieben. Sie beinhaltet die Losung der Starke mit einer thermostabilen a-Amylase, gefolgt von der vollstandigen Hydrolyse mit einer Amyloglucosidase. Die Glucose wird dann durch das Hexokinase/Glucose-6-phosphat-dehydrogenase-Verfahren bestimmt. Durch die Verwendung einer a-Amylase bei hoher Temperatur wird ein Autoklav zur Verkleisterung und Losung der Starke vor der Amyloglucosidasebehandlung nicht benotigt. Die a-Amylase wird der Probe vor der Erhitzung zugesetzt, und der Abbau beginnt, sobald die Verkleisterungstemperatur erreicht ist, bevor Retrogradation eintreten kann. Das Verfahren war denen mit Saure-oder Alkalibehandlung iiberlegen, Verfahren, die ebenfalls keinen Autoklaven erfordern.

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Cereal Chem. 74(4):497-501A variety of Rapid-Visco Analyser (RVA) operating conditions have been tested with starch, flour, and wholemeal for predicting the quality of wheats for the manufacture of Japanese white-salted noodles. Using starch as the substrate, an initial temperature of 60°C has been found to be optimum, and the best heating time from this initial temperature to the peak temperature of 95°C was ≈6 min. Significant correlations were found between peak viscosity of starch pastes and noodle quality under these operating conditions. For flour and wholemeal samples, the correlations were not as high as for isolated starch. The correlations with wholemeal or flour and noodle quality could be improved by the addition of α-amylase inhibitors. Measuring RVA viscosity of flour or wholemeal in the presence of silver nitrate gave viscosities which showed highly significant correlations with noodle quality. These correlations were similar to those obtained with isolated starch. It appears that the improvement is due to inhibition of the α-amylase present in grain and flour. Correlations were also observed between flour paste viscosity and alkaline noodle quality. These could be increased either by inhibiting α-amylase with silver nitrate or by pH adjustment with sodium carbonate but the change was not significant. The improvement of the correlations by α-amylase inhibitors in this sample set was not as great as observed with Japanese white noodles.

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