Exotic Corn Lines with Increased Resistant Starch and Impact on Starch Thermal Characteristics

Ten parent corn lines, including four mutants (dull sugary2, amylose‐extender sugary2, amylose‐extender dull, and an amylose‐extender with introgressed Guatemalen germplasm [GUAT ae]) and six lines with introgressed exotic germplasm backgrounds, were crossed with each other to create 20 progeny crosses to increase resistant starch (RS) as a dietary fiber in corn starch and to provide materials for thermal evaluation. The resistant starch 2 (RS2) values from the 10 parent lines were 18.3–52.2% and the values from the 20 progeny crosses were 16.6–34.0%. The %RS2 of parents was not additive in the offspring but greater RS2 in parents was correlated to greater RS2 in the progeny crosses (r = 0.63). Differential scanning calorimetry (DSC) measured starch thermal characteristics, revealing positive correlations of peak gelatinization temperature and change in enthalpy with %RS2 (r = 0.65 and r = 0.67, P ≤ 0.05); however, % retrogradation (a measure of RS3) and retrogradation parameters did not correlate with %RS2. The %RS2 and onset temperature increased with the addition of the ae gene, likely because RS delays gelatinization.     

Resistance to α‐Amylase Digestion in Four Native High‐Amylose Maize Starches

Native and processed high‐amylose maize starch (HAMS) is an important source of resistant starch (RS). The objectives of this work were to use an in vitro procedure to estimate the RS content of native granules from a series of ae‐containing HAMS genotypes, and to examine the nature of the α‐amylase resistant starch (ARS). By the method of Englyst et al (1992), RS for ae V, ae VII, ae su2, and ae du were estimated to be 66.0, 69.5, 69.5, and 40.6%, respectively. By transmission electron microscopy, most of the residual granules from ae V, ae VII, and ae su2 showed little evidence of digestion. Partially digested granules had a radial digestion pattern in the interior and an enzyme‐resistant layer near the surface. Size and chain‐length profile of constituents of ARS were similar to those of the native HAMS (unlike type 3 RS), consistent with complete hydrolysis in susceptible granule regions. Between crossed polarizers, many iodine‐stained native and residual HAMS granules had blue centers and pink exteriors, which may be due to a difference in orientation of the amylose‐iodine complexes in the exterior. Four granule color types were observed for ae du, differing in enzyme resistance. The high‐enzyme resistance of native HAMS granules may result from altered granule organization, which appears to vary among and within granules from ae‐containing genotypes.     

Classification of Single‐ and Double‐Mutant Corn Endosperm Genotypes by Near‐Infrared Transmittance Spectroscopy

A total of 1,176 grain samples representing 10 different single‐ and double‐mutant genotypic classes of specialty starch corn were used for developing various classification models based on near‐infrared transmittance spectra. The genotypes used included amylose‐extender (ae), dull (du), sugary‐2 (su2), waxy (wx), ae wx, ae du, ae su2, du wx and du su2. Two‐class classification models (only two genotypes compared) were developed using partial least squares analysis (PLS) while three‐way and multiclass models were examined using principal component analysis (PCA). The effectiveness of the calibrations was evaluated by examining the percentage of unknown grain samples incorrectly classified. In general, two‐class models performed better than multiclass models. However, they did not show improvement when discriminating among genotypes with overlapping amylose contents such ae du vs. ae and ae su2 vs. ae. Three‐way models including double‐mutants and their corresponding single‐mutant counterparts had misclassification percentages typically <5% using 14 PCA factors but again, with the exception of models including genotypes with overlapping amylose contents such as ae du vs. ae vs. du. The best multiclass model using all 10 genotypic classes simultaneously revealed only two classes (ae su2 and du) with misclassification rates >10% based on 16 PCA factors. This study demonstrates that, depending on the material to be considered, near‐infrared transmittance spectroscopy could be useful when segregation of specialty starch hybrids grain from other grain types is necessary.     

Comparison of Amylose Determination Methods and the Development of a Dual Wavelength Iodine Binding Technique

It has long been recognized that limitations exist in the analytical methodology for amylose determination. This study was conducted to evaluate various amylose determination methods. Purified amylose and amylopectin fractions were obtained from corn, rice, wheat, and potato and then mixed in proportion to make 10, 20, 30, 50, and 80% amylose content starch samples for each source. These samples, considered amylose standards, were analyzed using differential scanning calorimetry (DSC), high-performance size-exclusion chromatography (HPSEC), and iodine binding procedures to generate standard curves for each of the methods. A single DSC standard equation for cereal starches was developed. The standard curve of potato starch was significantly different. Amylose standard curves prepared using the iodine binding method were also similar for the cereal starches, but different for potato starch. An iodine binding procedure using wavelengths at 620 nm and 510 nm increased the precision of the method. When HPSEC was used to determine % amylose, calculations based on dividing the injected starch mass by amylose peak mass, rather than calculations based on the apparent amylose/amylopectin ratio, decreased the inaccuracies associated with sample dispersion and made the generation of a cereal amylose standard curve possible. Amylose contents of pure starch, starch mixtures from different sources with different amylose ranges, and tortillas were measured using DSC, HPSEC, iodine binding, and the Megazyme amylose/amylopectin kit. All the methods were reproducible (±3.0%). Amylose contents measured by these methods were significantly different (P < 0.05). Amylose measurements using iodine binding, DSC, and Megazyme procedures were highly correlated (correlation coefficient >0.95). DSC and traditional iodine binding procedures likely overestimated true amylose contents as residual butanol in the amylose standards caused interference. The modified two-wavelength iodine binding procedure seemed to be the most precise and generally applicable method. Each amylose determination method has its benefits and limitations.     

Structural Properties of Starch Fractions Isolated from Normal and Mutant Corn Genotypes Using Different Methods

The objectives of this research study were to isolate, evaluate, and compare the fine structures of starch fractions obtained from a wild‐type (normal) corn starch and amylose‐extender25, dull39, sugary2, and sugary1 corn mutants in the same genetic background using three different fractionation procedures based on gel‐permeation chromatography or alcohol‐precipitation methods. Starch fractions obtained from each of the three methods were enzymatically debranched and analyzed using high‐performance anion‐exchange chromatography with a postcolumn amyloglucosidase reactor and a pulsed amperometric detector. The separations were performed by fractionation on a GPC column, by precipitation with 1‐butanol, and by preferential precipitation with 1‐butanol and isoamyl alcohol. Using any of these methods, no apparent differences in the molecular weight distributions of amylopectin or of amylose among the different starches were observed. The proportions of branch chain lengths of the starch components obtained by the various fractionation methods were very similar among methods for each of the starch types analyzed, such as the predominance of long branch chains in ae25 corn and that of the short branch chains in su2 corn. Overall, the effect of the corn mutations was more important to the differences observed among the starch types than was the method of fractionation used.     

Production of Boiling‐Stable Granular Resistant Starch by Partial Acid Hydrolysis and Hydrothermal Treatments of High‐Amylose Maize Starch

The purpose of the present work was to examine whether partial acid hydrolysis (PAH) of a high‐amylose maize starch (ae‐VII) would enhance the effects of hydrothermal treatments to produce granular resistant starch (RS) that is stable to further heat treatment at atmospheric pressure. PAH ae‐VII starches were prepared by heating 35% (w/v) suspensions with 1% (w/w) HCl at 25°C for 6, 30, and 78 hr. Native and PAH starches were then treated by annealing (ANN) or heat‐moisture treatment (HMT). ANN was done at 70% moisture at 50, 60, or 70°C for 24 hr, and HMT was done at 30% moisture at 100, 120, or 140°C for 80 min. RS that survives boiling during analysis was determined by a modification of the AOAC method for determining total dietary fiber. RS was also determined by the Englyst method. Little change in the gelatinization enthalpy was found for ae‐VII starch after PAH, ANN, or HMT as individual treatments. After PAH, either ANN or HMT led to decreased gelatinization enthalpy. HMT and ANN alone increased boiling‐stable RS but decreased total RS. After PAH of ae‐VII, either ANN or HMT tended to increase the yield of boiling‐stable granular RS, with the greatest yield (≤63.2%) observed for HMT.     

Amylose and Amylopectin Interact in Retrogradation of Dispersed High-Amylose Starches

Retrogradation of three high-amylose starches (HAS: ae du, ae V, and ae VII) and common corn starch (CCS) was examined by dynamic oscillatory rheometry (7.5% [w/w] starch in 20% [v/v] dimethyl sulfoxide [DMSO]), differential scanning calorimetry (DSC; 30% [w/w] starch in water), and turbidity (0.5% [w/w] starch in 20% [v/v] DMSO). Nongranular lipid-free starch and starch fractions (amylose [AM], amylopectin [AP], and intermediate material [IM]) were studied. Gels were prepared by dispersing starches or fractions in 90% DMSO and diluting with water, followed by storage for seven days at 4°C. For AM from each starch, the elastic modulus (G′) was similar when heated from 6 to 70°C. The G′ of HAS AP gels at 6°C was higher than for CCS AP gels. For nongranular CCS and ae du gels, G′ dropped dramatically (≈100×) when heated from 6 to 70°C, less (≈10×) for ae V gels, and least (≈5×) for ae VII gels. By DSC, each AM endotherm had a peak temperature of ≈140°C, whereas all AP endotherms were complete before 120°C. Endotherms >120°C were not observed for any nongranular starch despite the high AM content of some starches. After cooling starch suspensions from room temperature to 5°C and subsequent rewarming to room temperature, each AM and the ae VII nongranular starch remained highly turbid. Each AP and the remaining nongranular starches lost turbidity during rewarming. Our work suggests that branched molecules of CCS and HAS influence gel properties of nongranular starches by inhibiting or altering AM-AM interactions.     

Wet‐Milling Characteristics of 10 Lines from Germplasm Enhancement of Maize Project Compared with Five Corn Belt Lines

The use of corn (Zea mays L.) hybrids with high grain yield and starch extractability has steadily increased in the processing industry. In light of widespread corn seed industry participation in the Germplasm Enhancement of Maize Project (GEM), which seeks to enhance exotic germplasm, future hybrids may contain more exotic sources in genetic backgrounds. It is necessary to establish and monitor physical, compositional, and milling characteristics of the new exotic breeding materials to determine the processing value. The present study was conducted to determine the wet‐milling characteristics of a set of GEM lines compared with typical Corn Belt lines. Ten GEM lines introgressed with exotic materials from Argentina, Chile, Cuba, Florida, and Uruguay and previously identified as having different starch yields, three commercial inbred lines, and two public inbred lines (B73 and Mo17) were analyzed using both near‐infrared transmittance (NIT) and a 100‐g wet‐milling procedure. There were statistical differences (P < 0.05) in the yield of wet‐milled fractions (starch, fiber, gluten, and germ). The GEM lines AR16035:S19‐227‐1‐B and CUBA117:S1520‐562‐1‐B had similar or better starch yield and starch recovery than B73 and the other adapted inbred lines, indicating that they may be useful in improving the proportion of extractable starch present in kernels of hybrids. Residual protein levels in the starch and gluten fractions were 0.26–0.32% and 38–45%, respectively. The starch yield of GEM lines from wet milling correlated positively with starch content from NIT and was negatively correlated with protein content of the corn kernels. Oil content in the germ varied from 50 to 60%. Our results indicate that incorporating GEM lines in a breeding program can maintain or even improve wet‐milling characteristics of Corn Belt materials if lines with appropriate traits are used.     

Physical Properties and Enzymatic Digestibility of Phosphorylated ae,wx, and Normal Maize Starch Prepared at Different pH Levels

Phosphorylated starches were prepared with sodium tripolyphosphate (STPP) at pH 6, 8, and 10 from waxy (wx, 3.3% amylose), normal (22.4% amylose), and two high-amylose (ae, 47 and 66% amylose) maize starches. After phosphorylation, the gelatinization peak temperature (T_p) decreased and pasting peak viscosity (PV) increased for all the starches except wx, which showed a slight increase in gelatinization temperature. There was a substantial effect of phosphorylation pH on paste viscosity. More crosslinking was found in ae starches with phosphorylation at pH 10. Sodium ions apparently decreased PV of all the phosphorylated starches while only slightly affecting PV of native starches. Phosphorylation increased swelling power of some of the starches, with maximum swelling power at phosphorylation pH 8 and minimum at pH 10. Maximum swelling power for wx starch after preparation was at pH 8 and minimum at pH 6. After phosphorylation, the clarity and freeze-thaw stability of all the starches was greatly increased compared with the native starches. Phosphorylation increased digestibility of ae starches but had little effect on wx and normal starches. After phosphorylation, the adhesiveness, springiness, and cohesiveness of all starch gels generally increased, the hardness of 47% ae and wx starches increased, and that of normal starches decreased. Enthalpy of gelatinization decreased after phosphorylation with the greatest decrease observed for ae starches. When the phosphorylation pH increased from 6 to 10, the brightness (L*) of all the phosphorylated starches decreased, while a* and b* of all the phosphorylated starch increased. Scanning electron micrographs showed some erosion on the surface of starch granules after phosphorylation.     

Gelatinization Properties of Starches from Three Successive Generations of Six Exotic Corn Lines Grown in Two Locations

The objectives of this research were to evaluate the intra‐ and interpopulation variability in gelatinization properties of starches from exotic corn lines and their derivatives when grown 1) during two successive years in the same location; and 2) in both temperate and tropical environments. Six novel exotic corn lines (two 100% exotic and four 25% exotic derived from a breeding cross developed by crossing an exotic hybrid with Corn Belt lines) were selected for this research because their starches have significantly different (and potentially useful) thermal properties from those found in starch from normal Corn Belt corn. The S_n (n = 3 for 25% exotic lines and n = 1 for 100% exotic lines) generations of the six exotic lines were self‐pollinated and grown in the winter nursery in Puerto Rico. Two successive generations (S_n+1 and S_n+2) of lines selected for low onset of gelatinization temperature were self‐pollinated and grown in the same environment near Ames, IA. To evaluate the effect of environment, the S_n+2 generation also was self‐pollinated and grown in the winter nursery in Puerto Rico. Thermal properties of starches from 10 single kernels from each line were analyzed by using differential scanning calorimetry (DSC) at a ratio of 4 mg of dry starch to 8 mg of distilled water. After subsequent generations, the differences in DSC gelatinization properties between selected kernels within each progeny line narrowed, suggesting increased homogeneity of starch structural properties within each line. Unusual thermal properties were fixed in some progeny lines. Environmental factors also affected the thermal properties of starch and a significant interaction between environment and genotype was observed. These results suggest that introgression of adapted germplasm with useful genes from exotic corn would increase the available genetic variability for starch functionality and allow the development of hybrids with important value‐added traits.     

Thermal Properties of Corn Starch Extraction Intermediates by Differential Scanning Calorimetry

Thermal properties of corn starch extraction intermediates from four types of corn were studied using differential scanning calorimetry. Starch at four different stages of extraction, including a standard single-kernel starch isolation procedure and three starch extraction intermediates, was isolated from mature corn kernels of B73 and Oh43 inbreds and the mutants of waxy (wx) and amylose extender (ae) in an Oh43 background. Differences in thermal properties and moisture and protein contents of starch from the extraction stages were statistically analyzed. Most thermal properties (gelatinization and retrogradation onset temperatures, gelatinization and retrogradation ranges, gelatinization and retrogradation peak temperatures, gelatinization and retrogradation enthalpies, peak height index, and percentage of retrogradation) of starches extracted at stage 3 intermediate (a procedure that did not include a final washing step) were similar to those of starch extracted by the standard single-kernel isolation procedure. Values for gelatinization peak temperature, gelatinization enthalpy, and peak height index were different between the standard and the stage 3 intermediate. The values obtained from starches extracted at stage 3, however, were consistent and predictable, suggesting that this extraction intermediate might be used in screening programs in which many starch samples are evaluated. By using the stage 3 extraction, samples could be evaluated in three rather than four days and the procedure saved ≈0.5 hr of labor time. The other two starch extraction intermediates, which excluded filtering and washing or filtering, washing, and steeping, produced starch with thermal properties generally significantly different from starch extracted by the standard single-kernel isolation procedure.     

Structural Changes of Debranched Corn Starch by Aqueous Heating and Stirring

Aqueous dispersions (2 mg/mL) of debranched corn starches of different amylose contents (waxy, normal, and high‐amylose) were subjected to extensive autoclaving and boiling‐stirring, and then the changes in starch chain profile were examined using medium‐pressure, aqueous, size‐exclusion column chromatography. As autoclaving time increased from 15 to 60 min, weight‐average chain length (CL_w) of waxy, normal, and high‐amylose corn starches determined using pullulan standards decreased from 46 to 41.2, from 122.1 to 96.3, and from 207.3 to 151.8, respectively. Number‐average chain length (CL_n) measured by the Nelson‐Somogyi method also decreased from 23.0 to 18.4, from 26.4 to 21.8, and from 66.5 to 41.5, respectively, indicating that thermal degradation of starch chains occurred. The CL_w/CL_n ratio for normal corn starch was higher than that for waxy corn starch, indicating an increase in polydispersity of the amylose fraction. Thermal degradation was also observed when the debranched starch was subjected to the boiling‐stirring treatment (0–96 hr). During 96 hr, the CL_w and relative proportion of B≥2 chains of amylopectin released by debranching waxy corn starch increased, whereas those of B1 chains decreased. This change may indicate physical aggregation of B1 chains. But branches from normal and high‐amylose corn starches showed increases in CL_w and the proportion of both B1 and B≥2 chains, along with substantial decreases in those of amylose chains. Therefore, thermal degradation of amylose was greater than that of amylopectin.     

Selecting and Sorting Waxy Wheat Kernels Using Near‐Infrared Spectroscopy

An automated single kernel near‐infrared (NIR) sorting system was used to separate single wheat (Triticum aestivum L.) kernels with amylose‐free (waxy) starch from reduced‐amylose (partial waxy) or wild‐type wheat kernels. Waxy kernels of hexaploid wheat are null for the granule‐bound starch synthase alleles at all three Wx gene loci; partial waxy kernels have at least one null and one functional allele. Wild‐type kernels have three functional alleles. Our results demonstrate that automated single kernel NIR technology can be used to select waxy kernels from segregating breeding lines or to purify advanced breeding lines for the low‐amylose kernel trait. Calibrations based on either amylose content or the waxy trait performed similarly. Also, a calibration developed using the amylose content of waxy, partial waxy, and wild‐type durum (T. turgidum L. var durum) wheat enabled adequate sorting for hard red winter and hard red spring wheat with no modifications. Regression coefficients indicated that absorption by starch in the NIR region contributed to the classification models. Single kernel NIR technology offers significant benefits to breeding programs that are developing wheat with amylose‐free starches.     

Effects of Amylose,Corn Protein,and Corn Fiber Contents on Production of Ethanol from Starch‐Rich Media

The effects of amylose, protein, and fiber contents on ethanol yields were evaluated using artificially formulated media made from commercial corn starches with different contents of amylose, corn protein, and corn fiber, as well as media made from different cereal sources including corn, sorghum, and wheat with different amylose contents. Second‐order response‐surface regression models were used to study the effects and interactions of amylose, protein, and fiber contents on ethanol yield and conversion efficiency. The results showed that the amylose content of starches had a significant (P < 0.001) effect on ethanol conversion efficiency. No significant effect of protein content on ethanol production was observed. Fiber did not show a significant effect on ethanol fermentation either. Conversion efficiencies increased as the amylose content decreased, especially when the amylose content was >35%. The reduced quadratic model fits the conversion efficiency data better than the full quadratic model does. Fermentation tests on mashes made from corn, sorghum, and wheat samples with different amylose contents confirmed the adverse effect of amylose content on fermentation efficiency. High‐temperature cooking with agitation significantly increased the conversion efficiencies on mashes made from high‐amylose (35–70%) ground corn and starches. A cooking temperature of ≥160°C was needed on high‐amylose corn and starches to obtain a conversion efficiency equal to that of normal corn and starch.     

Restriction of Starch Granule Swelling by Iodine During Heating

Differences in the degree of starch granule swelling and granule morphology are shown as a function of iodine concentration during heating. The observations reveal a restriction and delay in wheat and corn starch swelling in presence of iodine (0.02%) and a lack of swelling at higher iodine concentration (0.2%). The presence of iodine during heating did not influence waxy corn starch granule swelling, even at the higher concentration. A delay in the increase in paste viscosity during heating was observed, and gel formation was precluded after cooling at the higher iodine concentration. Waxy corn starch pastes formed a weak gel even at the higher iodine concentration. Spectrophotometric analyses showed that polymers leach into the solution when heated in the presence of 0.02% iodine, while no leaching was observed at 0.2% iodine concentration. Furthermore, the length and the amount of polymers leaching from normal corn were different from that observed for wheat starch. Thermal analyses reveal a shift in the onset of gelatinization temperature and an increase in the enthalpy in the amylose‐lipid region of the endotherm. While the iodine‐polymer complex did not appear to exhibit an endotherm during heating, the granules exhibited a diffused polarized cross, suggesting the presence of an ordered complex.     

Effects of Resistant Starch and Fiber from High‐Amylose Non‐Floury Corn on Tortilla Texture

A high‐amylose, non‐floury corn, a floury corn, and a 1:1 blend were made into masa and then tortillas. The masa flour made with the high‐amylose corn had a greater amount of resistant starch (RS 28.8%) and a greater amount of total dietary fiber (TDF 42.1%) than that with the floury corn (RS 2.9%, TDF 9.6%), producing a high‐fiber tortilla. The masa was evaluated for pasting properties using a Rapid ViscoAnalyser (RVA). The high‐amylose masa slurry gelatinized little at 95°C. The floury masa had the greatest peak viscosity, whereas the 1:1 blend was intermediate in value. Sensory evaluations of the tortillas for the textural attributes showed the floury tortillas to be chewier, more rollable, and grittier than the high‐amylose tortillas, whereas the blend tortillas were intermediate for most attributes. The cutting force of the high‐amylose tortillas, measured on a texture analyzer, was very low; the blend and floury tortillas required more force. Chewiness was correlated to rollability (r = 0.99, P = 0.05). The %RS and %TDF were correlated to rollability (r = –0.99), and cutting force (r = 0.99). The floury and blend tortillas had firm textures expected of desirable tortillas, whereas the high‐amylose tortillas broke under little force, and would not roll. The high‐amylose tortillas had high amounts of RS and TDF but poor texture. The blend tortillas retained most floury tortilla textural properties, making them suitable products for consumer use.     

Characterization of Starch in Aegilops Species

Starch was extracted and cleaned from 99 accessions of 20 species of Aegilops and also from 200 accessions of hexaploid wheat. Amylose content was determined by iodine staining and absorbance at 535 and 620 nm. Particle‐size distribution was determined by laser scattering. The amylose content of the Aegilops accessions did not exceed the extremes found in domesticated wheat. Aegilops species, on the whole, had a lower content of small particles than the hexaploid wheats. There was no correlation between amylose content and particle‐size distribution. Some species of Aegilops may be useful sources of low‐starch B‐type granules for hexaploid wheat, if the trait can be transferred, but they are unlikely to contribute to further variation in amylose content.     

Novel ssIIa Alleles Produce Specific Seed Amylose Levels in Hexaploid Wheat

Novel starch synthase II (ssIIa ) alleles were created in a soft wheat (Triticum aestivum L.) via ethyl methanesulfonate mutagenesis and combined by crossing to create four unique ssIIa triple mutant haplotypes. A range of starch amylose content was obtained from the ssIIa triple mutant lines (35.6–46.8%), with moderate increases in amylose content found in lines carrying two ssIIa null alleles and one partially functional ssIIa allele. Seeds from all ssIIa triple mutants had significantly higher amylose and protein contents but also lower starch content, kernel weight, and flour swelling power compared with their wild‐type sister lines. Seed starch amylose content was negatively correlated with individual kernel weight (r = –0.54), starch content (r = –0.85), and flour swelling power (r = –0.91) but positively correlated with grain protein (r = 0.78), demonstrating that unique ssIIa triple mutant null combinations confer defined levels of seed starch amylose, protein content, and kernel size. The ssIIa mutant lines also had a hard grain texture (≈86 single kernel characterization system units), whereas all parental material and wild‐type sister lines had soft grain texture (≈35). This change in grain texture was independent of Ha locus genotype, because all lines carried a functional (soft type) Ha locus. The ssIIa alleles and allelic combinations reported here demonstrate the ability to create defined levels of seed starch, amylose, and protein.     

Properties of Starches from Near‐Isogenic Wheat Lines with Different Wx Protein Deficiencies

To determine the effect of amylose content on the starch properties, the amylose content, pasting properties, swelling power, enzymatic digestibility, and thermal properties of partial and perfect waxy types along with their wild‐type parent were analyzed. As expected, amylose content decreases differently in response to the loss of each Wx gene, showing the least response to Wx‐A1a. Most of the characteristics, except the thermal properties of the amylose‐lipid complex in differential scanning calorimetry (DSC), differed significantly among the tested types. Furthermore, the breakdown, setback, and pasting temperatures from the Rapid Visco Analyser (RVA) and the enzymatic digestibility, swelling power, peak temperature, and enthalpy of starch gelatinization from DSC showed a correlation with the amylose content. The relationships between the peak viscosity from the RVA and the onset temperature of starch gelatinization determined by DSC with amylose content of the tested materials were not clear. Waxy starch, which has no amylose, showed a contrasting behavior in starch gelatinization compared with nonwaxy starches. Among the nonwaxy starches, lower setback, lower pasting temperature, higher enzyme digestibility, higher peak temperature, higher enthalpy of starch gelatinization, and higher swelling were generally associated with low amylose starches.     

Interactions Among Wx Genes and Effects of a Mutated Wx‐D1e Allele on Starch Properties in Wheat

The waxy mutant wheat Tanikei A6599‐4 contains little amylose and exhibits a stable hot paste viscosity. It has null alleles at the Wx‐A1 and Wx‐B1 loci and a mutated allele at the Wx‐D1 locus (Wx‐D1e). From the cross‐combination of Kanto 123 (normal amylose line) and Tanikei A6599‐4, 51 DH (doubled haploid) lines were produced and their genotypes were determined by SDS‐PAGE and a derived cleaved amplified polymorphic sequence (dCAPS) analysis. Eight genotypes were obtained at the expected ratio. Using a Rapid Visco Analyser (RVA), all the lines with the same genotype as Tanikei A6599‐4 showed a stable hot paste viscosity. The other lines did not show a stable hot paste viscosity regardless of the presence of the Wx‐D1e allele. When two genotypes with the same Wx‐A1 and Wx‐B1 alleles were compared, the one with Wx‐D1e contained less amylose and exhibited a lower final viscosity and a lower setback with RVA. Although the Wx‐D1e allele produces an almost inactive Wx protein, these findings suggest that this allele contributes at some level to starch synthesis.