Guidelines for in-season nitrogen application for maize (Zea mays L.) based on soil and terrain properties

Nitrogen (N) fertilizers are often applied to maize (Zea mays L.) in excess of economically optimal rates because of the uncertainty of dealing with seasonal and spatial variability. A better understanding of the relationships among field, apparent soil electrical conductivity (EC_a), elevation, slope and seasonal characteristics is therefore essential for performing optimal variable-rate N applications. This study focused on responses during the exponential growth phase, when it is critical that N supply be not limited. Measurements at high spatial resolution allowed to understand the effects of the relationships among N, EC_a, elevation, slope and season on future yield formation. The study was conducted over three years (2005-2007). Mid-season growth responses to applied N were greatest where EC_a levels were high and elevation was low in 2005 and 2007, but not in 2006. Areas with slope ≥1 degree were also more responsive to N rates. Overall best mid-season growth was found in areas of low EC_a, high Elevation and low Slope. However, the best responses to in-season N fertilization were found in areas with opposite properties (high EC_a, low Elevation and high Slope). Indeed, relatively high rates of in-season N were needed to enhance crop growth in areas of high EC_a, low Elevation and high Slope, which are characteristic of unfavourable growth conditions. In counterpart, lower N rates were sufficient for optimal growth in soils at low EC_a high Elevation and low Slope. Also, despite the fact that conditions of high soil variability were specifically selected for the study, the effects and interactions reported for soil NO₃-N content were small. The interaction of EC_a with early seasonal precipitation is likely a key relationship to consider in variable-rate N application: low-lying areas with fine soil texture showed the greatest dependence on weather for optimal N rates. Indeed, the relationships among factors influencing the response to in-season N fertilization were stronger when seasonal conditions were particularly favourable to maize growth. These results are fundamental to the establishment of in-season application rules for spatially variable N algorithms.     

Variability of grain productivity and energy profile of maize (Zea mays L.) genotypes

The goal of the present work was to test if there is genetic variability between precocious and superprecocious cycle maize genotypes in terms of grain productivity and energy profile. Data from two separate experiments were used. An experiment was conducted with 36 precocious cycle genotypes and another experiment was conducted with 22 superprecocious cycle genotypes. The following variables were measured: grain productivity (PROD), ethereal extract (EE), starch (ST), amylose (AML), and nitrogen-corrected apparent metabolizable energy (AMEn). For all variables, basic statistics and analysis of variance was performed. For each experiment, the genotypic correlation matrix was calculated and the multicollinearity was evaluated. Then, the Mahalanobis generalized distance matrix was calculated, the clustering of the genotypes was performed and the averages of the groups formed were compared, separately for each experiment. There was genetic divergence between precocious genotypes, as well as between superprecocious genotypes, especially in terms of grain PROD, EE concentration, and AML concentration. Groups of more productive genotypes presented lower EE and higher AML grain concentrations. The results of the present study indicated that it is possible to plan crossings between groups of genotypes in terms of PROD, EE and AML, with the goal of maximizing heterosis.     

Adoption of leaf color chart for nitrogen use efficiency in rice: Impact assessment of a farmer-participatory experiment in West Bengal,India

Unbalanced and excessive use of N-fertilizers causes environmental pollution, lodging of plants and increased pest pressure, in addition to increased cost to farmers from excessively applied fertilizers and pesticides. N application at the right time and in right amount is critical for healthy plant and environment. Rice leaf color intensity is directly related to leaf chlorophyll content and leaf nitrogen status. The concept for the use of leaf color as an indicator to apply N in rice was crystallized during 1990s. The International Rice Research Institute and the Philippine Rice Research Institute developed a leaf color chart (LCC) that helps guide farmers for real-time nitrogen management in rice farming. The technology is inexpensive, and easily affordable by most resource poor rice farmers. In 2003 we initiated a farmer-participatory research to validate real-time N management in rice by the use of LCC in West Bengal state of India. After 3 years of validation research, a survey was conducted to assess the adoption and impact of LCC. The survey was conducted in both intervention and adjacent control villages and data were collected from 20% farm households selected randomly. In this paper, we report findings of the study on the determinants of adoption of LCC, and its effect on fertilizer and pesticides use.     

Control of volunteer cereals with post-emergence herbicides in maize (Zea mays L.)

Volunteer winter cereals are found sporadically in maize (Zea mays L.) fields across southern Ontario. Seven field trials were conducted over a two-year period (2006 and 2007) at four locations to determine the efficacy of five acetolactate synthase (ALS)-inhibiting herbicides for the control of volunteer cereals applied at two post-emergence application timings (2–4 and 4–7 maize leaf tips). The volunteer cereals were a hard red winter wheat (Triticum aestivum L.) (‘Hyland AC Morley’), soft red winter wheat (‘Pioneer 25R47’), soft white winter wheat (‘Pioneer 25W41’), and a autumn rye (Secale cereale L.) (‘FR’) cultivar. Volunteer cereal competition in maize resulted in a yield reduction of up to 44%. Foramsulfuron, nicosulfuron, nicosulfuron/rimsulfuron provided greater than 70% control of the volunteer cereals at 56 days after treatment (DAT), while primisulfuron and rimsulfuron provided greater than 60% control. Volunteer cereal control with early and late application was greater than 82 and 61%, respectively. Hard red winter wheat was the most sensitive to the ALS-inhibiting herbicides with control of 84–93%. Soft red and soft white winter wheat cultivars were intermediate in sensitivity with control of 76–87%, while autumn rye was the least sensitive with control of at 56–71% control at 56 DAT. Maize yields were improved when volunteer cereals were controlled with the use of herbicides compared to the weedy control, but were lower than the weed-free control. Early herbicide application resulted in improved control of volunteer cereals and higher maize yield.     

Conversion into bioethanol of insect (Sitophilus zeamais Motschulsky), mold (Aspergillus flavus Link) and sprout-damaged maize (Zea mays L.) and sorghum (Sorghum bicolor L. Moench)

The bioconversion into ethanol of insect (Sitophilus zeamais), mold (Aspergillus flavus) and sprout-damaged maize and sorghum was investigated. Kernel test weight losses due to insect damage in maize were almost twice compared to sorghum (18.6 vs. 10.7%). All damaged kernels lost some of the starch and increased soluble sugars, ash and crude fiber. The mold-damaged sorghum contained approximately five times more FAN compared to the control. The sprout-damaged kernels contained the highest amounts of reducing sugars prior (11 g/L) to and at the end (146.5 g/L) of liquefaction with α-amylase. Ethanol yields based on the already damaged grain indicated that sprout-damaged kernels yielded similar amounts compared to sound kernels (381.1 vs. 382.6 L/ton and 376.6 vs. 374.8 L/ton of sorghum or maize respectively). The insect-damaged maize and sorghum have reduced ethanol yields compared with the controls (29 and 23% respectively), and this negative result was mainly due to dry matter losses during the inadequate storage. Despite differences in ethanol yield, all treatments have similar conversion efficiencies (76.1–89.9%) indicating the robustness of yeast facing biotic-damaged feedstocks. This research demonstrates that the use of already damaged insect, mold or sprouted kernels is feasible and a good alternative for biorefineries.     

Protein quality and endosperm modification of quality protein maize (Zea mays L.) under two contrasting soil nitrogen environments

Quality protein maize (QPM) breeding involves the combined use of the opaque-2 (o2) gene and the genetic modifiers of the o2 locus to develop cultivars with modified kernel endosperm, and increased concentrations of lysine and tryptophan. This study was designed to assess grain yield performance, endosperm modification, and protein quality and quantity under two contrasting soil nitrogen environments. A 15-parent diallel cross was evaluated under one low nitrogen stress and one optimal nitrogen environment each at Harare (Zimbabwe) and Bako (Ethiopia). Most QPM hybrids showed higher protein quality levels than the best non-QPM check under both conditions. Protein concentration tended to vary across nitrogen levels, but not endosperm type. Significant differences were found for the test of main effect (nitrogen-level) for endosperm modification and tryptophan concentration. This indicated that QPM maintains quality even under low soil nitrogen, a widespread condition in Africa. General combining ability (GCA) mean squares were highly significant for most protein quality traits for each environment and across environments whereas specific combining ability (SCA) mean squares were not significant in most cases. This indicated that additive gene effects were primarily responsible for variation of most traits evaluated and hence progeny performance can adequately be predicted on the basis of parental performance. Inbred lines P2, P4 and P12 had desirable GCA effects for endosperm modification while P1 and P3 had the best GCA for tryptophan concentration in grain. The current study suggests that hybrids with desirable endosperm modification, protein quality and stable performance under low nitrogen stress and optimal conditions can be produced with careful selection.     

The usefulness of iron bioavailability as a target trait for breeding maize (Zea mays L.) with enhanced nutritional value

Iron (Fe) deficiency is the most widespread nutritional problem, affecting as many as half of the world's population. Only a small fraction (2-15%) of Fe from plant sources is typically bioavailable, that is, available for absorption and nutritionally useful for humans. This study evaluated Fe concentration and bioavailability for three diverse sets of 12, 14 and 16 maize hybrids grown in two- or three-location trials to assess the feasibility of selecting for Fe bioavailability in breeding programs. Bioavailability of Fe, assessed using the in vitro digestion/Caco-2 cell model, varied significantly among hybrids in two of the three trials. Location effects were larger than location by genotype interaction effects, additive but not non-additive gene action was significant, and heritability estimates were mostly between 0.55 and 0.65 for Fe bioavailability estimators. Bioavailability of Fe was not associated with Fe concentration in grain or with grain yield. Negative correlation of Fe bioavailability with zinc concentration in grain for one of the three hybrid trials, and positive correlation with provitamin A concentrations in one trial were indicative of inhibitor and enhancer effects on Fe bioavailability, respectively. Although use of the Caco-2 cell model is promising, particularly because it integrates the whole meal, or food matrix effect on Fe bioavailability, the complex nature of the assay and moderate heritability of bioavailability estimators make it most suitable as an intermediate selection tool, following high throughput selection for molecular markers of Fe bioavailability, currently in development by other researchers, and preceding validation and efficacy trials with animal and human models.     

Genotypic variation for root traits of maize (Zea mays L.) from the Purhepecha Plateau under contrasting phosphorus availability

Phosphorus (P) deficiency is a major constraint for maize production in many low-input agroecosystems. This study was conducted to evaluate genotypic variation in both root (root architecture and morphology, including root hairs) and plant growth traits associated with the adaptation of maize landraces to a P-deficient Andisol in two locations in the Central Mexican highlands. Two hundred and forty-two accessions from the Purhepecha Plateau, Michoacan were grown in Ponzomaran with low (23 kg P₂O₅ ha⁻¹) and high (97 kg P₂O₅ ha⁻¹) P fertilization under rain-fed field conditions, and subsequently a subset of 50 contrasting accessions were planted in the succeeding crop cycle in Bonilla. Accessions differed greatly in plant growth, root morphology and P efficiency defined as growth with suboptimal P availability. The accessions were divided into 3 categories of P efficiency using principal component and cluster analyses, and 4 categories according to the retained principal component and their relative weight for each genotype in combination with growth or yield potential. The distribution of accessions among three phosphorus efficiency classes was stable across locations. Phosphorus-efficient accessions had greater biomass, root to shoot ratio, nodal rooting, nodal root laterals, and nodal root hair density and length of nodal root main axis, and first-order laterals under P deficiency. Biomass allocation to roots, as quantified by the allometric partitioning coefficient (K) was not altered by P availability in the efficient accessions, but inefficient accessions had a lower K under low P conditions. Accessions with enhanced nodal rooting and laterals had greater growth under low P. Dense root hairs on nodal root main axes and first-order laterals conferred a marked benefit under low P, as evidenced by increased plant biomass. Late maturity improved growth and yield under low P. These results indicate that landraces of the Central Mexican highlands exhibit variation for several root traits that may be useful for genetic improvement of P efficiency in maize.     

Association between line per se and hybrid performance under excessive soil moisture stress in tropical maize (Zea mays L.)

Excessive moisture (EM) stress during the summer–rainy season is one of the major production constraints for maize (Zea mays L.) in large areas of South and South-East Asia. A key question in breeding for tolerance to excessive moisture is the extent to which the performance of maize hybrids can be predicted on the basis of per se performance of inbred lines under excessive moisture. We attempted to identify the relationship between morpho-physiological traits and grain yield measured on inbred parents and their single cross progenies under EM stress. Responses of various morpho-physiological traits, except days to 50% anthesis, differ significantly under normal versus EM stress. Superiority of hybrid progenies over parental inbred lines increased under EM stress, suggesting that hybrids were comparatively more tolerant to EM stress than inbred progenies. Across moisture regimes, all morpho-physiological traits of hybrids, except lodging and root porosity under normal moisture, were found to be positively and significantly correlated with mid-parent traits. Our data suggest that per se performance of lines was a relatively more important factor in determining hybrid performance under EM stress, while under optimum soil moisture conditions mid-parent heterosis was relatively more important than per se performance of mid-parent. Phenotypic correlation between hybrid and mid-parent yields showed a strong relationship under EM stress (r = 0.66^**). The relationship was statistically significant under normal moisture as well, though it was comparatively weak (r = 0.41^*). Our findings suggest that performance of hybrid progenies under excessive moisture can be predicted and improved to some extent on the basis of their inbred parents that have been systematically selected and improved for EM stress.     

Tolerance to excess moisture in maize (Zea mays L.): susceptible crop stages and identification of tolerant genotypes

Excess moisture (water-logging) during the summer–rainy season is one of the major production constraints for maize (Zea mays L.) in a large area of Southeast Asia. Identification and development of genotypes capable of withstanding the stress conditions could be an ideal and affordable approach suitable for resource poor maize-growing farmers of such areas. We attempted to identify the most susceptible/critical crop stage(s) of maize for excess moisture stress, and to develop a screening technique and selection strategies for identification of germplasm tolerant to excess moisture stress. Among the four crop stages, i.e. early seedling (V2), knee-high (V7), tasseling (VT) and milk stage (R1), V2 was found to be highly susceptible, followed by the V7 stage. A screening technique (cup method) was developed/standardized, and was found to be an efficient technique for large-scale screening of maize genotypes against excess soil moisture stress. Germplasm was screened using this technique followed by field evaluation at the V7 growth stage (seventh leaf visible). Excess soil moisture stress severely affected various growth and biochemical parameters, impaired anthesis and silking, and eventually resulted in poor kernel development and yield. However, remarkable variability was found among the genotypes studied. Genotypes with good carbohydrate accumulation in stem tissues, moderate stomatal conductance, <5 days ASI, high root porosity, and early brace root development ability have been found to have good tolerance against the hypoxia/anoxia caused by excess soil moisture conditions.     

Nitrogen use efficiency in selected rice (Oryza sativa L.) genotypes under different water regimes and nitrogen levels

Water and nutrient availability are two major constraints in most rice-based rainfed shallow lowland systems of Asia. Both stresses interact and contribute to the low productivity and widespread poverty in this environment. The objective of this study was to improve the understanding of interaction between the two factors and to identify varietal characteristics beneficial for productivity in a water- and nutrient-limited rice environment. For this purpose, we screened 19 rice genotypes adapted to different rice environments under two water and two nutrient treatments during the wet season of 2004 and 2005 in southern Luzon, Philippines. Across all genotypes tested and in comparison with the irrigated control, rainfed conditions reduced grain yield of the treatment without N application by 69% in 2004 and by 59% in 2005. The mean nitrogen fertilizer response was highest in the dryer season of 2004 and the rainfed treatment, indicating that water stress had no effect on fertilizer response. Nitrogen application reduced the relative yield loss to 49% of the irrigated treatment in 2004 and to 52% of the irrigated treatment in 2005. Internal efficiency of N (IEN) and recovery efficiency of applied N (REN) were significantly different between genotypes, but were not affected by water availability (REN) or by water and nutrient availability (IEN). In contrast, grain yield and total N uptake were affected by cultivar, N and water availability. Therefore, germplasm for rainfed environments should be screened under conditions of limited and good nitrogen and water supplies. The four best cultivars, CT6510-24-1-2, IR55423-01, IR72, and IR57514-PMI5-B-1-2, performed well across all treatments and both years. Except for IR72, they were all characterized by medium height, medium duration, high early vigor, and a moderate level of drought tolerance. This combination of characteristics seems to enable the optimal use of limited water and nutrient resources occurring in many shallow rainfed lowlands. We also concluded that moderate drought stress does not necessarily affect the response to moderate N rates, provided that drought does not induce high spikelet sterility and that fertilizer N is properly managed.     

Effect of nitrogen fertilizer on the growth and survival of Rhopalosiphum padi (L.) and Sitobion avenae (F.) (Homoptera: Aphididae) on different wheat cultivars

Increased nitrogen applications to crops influence plant-insect interactions and potentially increase herbivore population growth. The effect of nitrogen applications on the interactions between a wheat host plant, Triticum aestivum, and its herbivores, Sitobion avenae (Fabricius, 1775) and Rhopalosiphum padi (Linnaeus, 1758) was investigated. Adult body weight, fecundity, maturity rate and longevity of the aphids were measured under different nitrogen application on four wheat varieties, Solstice, Einstein, Deben and Alchemy. Adult body weight of R. padi on Alchemy and of S. avenae on Deben was lowest. Fecundity of R. padi was lesser on Deben and Alchemy while fecundity of S. avenae was least on Alchemy. Days to reach maturity were same on all varieties in S. avenae but in R. padi, Deben positively affected the days to reach maturity. Longevity of S. avenae and R. padi was the same on all varieties. N-fertilizer had a positive effect on the adult weight achieved by S. avenae and R. padi. Fecundity and longevity of both species were also positively correlated with N-fertilizer application. In R. padi, N-fertilizer reduced the time to reach maturity but not in S. avenae.     

Differences in leaf yield and indigo precursors production in woad (Isatis tinctoria L.) and Chinese woad (Isatis indigotica Fort.) genotypes

Isatis tinctoria L. (woad) is one of the earliest known sources of indigo in Europe where it was cultivated since the Middle Ages. Isatisindigotica Fort. (Chinese woad), widely distributed in China, had been used from ancient times as indigo-producing plant and medicinal plant. Both species produce indigo precursors indican (indoxyl β-d glucoside) and isatan B (indoxyl ketogluconate) in their leaves. In order to identify new suitable crops for indigo production in Italy, 17 woad lines were studied under field conditions in Central Italy (Pisa, 43°40′N, 10°19′E) from 2001 to 2003. We analyzed the effects of year, genotype, and harvest times together with their reciprocal interactions on leaf yield and indigo precursors production. Woad lines were then compared with seven I. indigotica lines in a field crop experiment set up in 2003. Extraction and quantification of indigo precursors were accomplished by HPLC-ELSD. Isatan B and indican content, as well as equivalent indigo and fresh/dry leaf yield, were compared between species and among genotypes.     

Evaluation of some weed control treatments for long season weed control in maize (Zea mays L.) under zero and minimum tillage at Samaru, in Nigeria

The effect of weed control treatments for long season control of weeds in maize under zero and minimum tillage was evaluated at Samara in northern Nigeria. Among the weed control treatments evaluated, soil ridging plus application of either 2,4-D or atrazine at 1.5 kg a.i/ha performed well as they effectively controlled weeds and resulted in better growth and a grain yield that was comparable to the hand weed control. Maize production under minimum tillage was better than under zero tillage.     

Nitrogen compounds in organs of two sugar beet genotypes (Beta vulgaris L.) during the season

A major aim in sugar beet breeding is to decrease the concentration of soluble N compounds in the beet to avoid negative impacts during beet processing. Due to its importance and analytical limitations the only selection criterion is amino N analyzed in the beet at harvest. It is thus not clear when genotypic differences are established during the season. Furthermore, it is not known whether selection for low amino N affects other N compounds in the beet and the N composition of other plant organs. Therefore, the concentrations of total N and soluble N compounds (protein, amino N, glutamine, betaine, nitrate) were investigated during the season in different organs (leaf blades, petioles, crown, beet) of two sugar beet genotypes differing in the amino N concentration of the beet. Field trials were carried out at three sites in 2002 and in 2003 with harvests at three times from 100 to 170 days after sowing.     

Impact of nitrogen amount and timing on the potential of acrylamide formation in winter wheat (Triticum aestivum L.)

Acrylamide (AA), a potential human carcinogen, is formed in strongly heated carbohydrate-rich food as a part of the Maillard reaction. The amino acid asparagine (Asn) and reducing sugars are considered to be the main precursors for AA formation. In a 2-year field trial the impact of nitrogen (N) amount and timing on the content of AA precursors and the potential of AA formation in different winter wheat cultivars (cv.) were studied in association with respective grain yields and parameters of baking quality. Depending on year, cultivar and nitrogen treatment Asn contents ranged between 4 and 18 mg 100 g⁻¹ flour dry-matter (DM). Nitrogen treatments affecting crude protein contents in flours above 13% caused a considerable increase in free Asn. Nitrogen amounts of 220 kg N ha⁻¹ increased the contents of free Asn by between 130% and 270% depending on year and cultivar compared to the untreated controls. A close linear correlation between the content of free Asn and the potential of AA formation (2004: R² = 0.89, 2005: R² = 0.83) could be observed, whereas no correlation could be found between reducing sugars and the potential of AA formation, pointing to the importance of free Asn as the limiting and thus determining factor for the AA formation potential in wheat flours. To reach high crude protein contents and good sedimentation values demanded for breadstuffs, nitrogen amounts of at least 180 kg N ha⁻¹ were necessary. Nitrogen fertilization measures resulting in high crude protein contents above 13% enhanced the potential of AA formation by increasing the content of free Asn in flours. As long as demands from traders and producers for flour with high crude protein contents are not revised, lowering Asn contents and thus the potential for AA formation by application of N amounts below 180 kg N ha⁻¹ and abandoning the late application of N do not appear to be successful ways to reduce the risk of AA formation in breadstuffs.     

Optimization of nitrogen rate and seed density for safflower (Carthamus tinctorius L.) production under low-input farming conditions in temperate climate

Field experiments under low-input farming conditions were conducted in South West Germany (lhinger Hof) and North Switzerland (Wil) in 2004 and 2005 aimed at optimizing nitrogen rate and seed density for the production of the newly introduced safflower (Carthamus tinctorius L.). The experiments were laid out in a four-replicated-split plot design with three nitrogen fertilizer rates (0, 40, 80 kg/ha) as main plots and cultivars (Sabina, Saffire, BS-62915) and seed densities (50, 100, 150 seeds/rn2) randomized in split plots. It was shown that many traits responded differentially across environments to rate of nitrogen and seed density. Application of 40 and 80 kg N/ha did not significantly affect most of the investigated traits. At Ihinger Hof, the total nitrogen fertilizer needed to maximize safflower yield was estimated to be 86 kg N/ha. At Wil, residual soil nitrogen alone resulted in satisfactory seed yield when safflower followed a crop fertilized at a commercial rate. The nitrogen rate × seed density interaction was only significant for seed yield and Alternaria leaf spot disease. Nitrogen rates provided significant increases in seed yield at high seed density compared to low seed density. Seed density did not reveal any significant variation in seed yield, oil content, and oil yield. On average, the low seed density produced substantially higher numbers of heads/plant and seeds/plant compared to medium and high densities. These results demonstrate the ability of safflower to use residual soil nitrogen efficiently and to compensate for low plant density.     

Polymorphisms in the ALS gene of weedy rice (Oryza sativa L.) accessions with differential tolerance to imazethapyr

Herbicide-resistant Clearfield™ rice technology allows the use of ALS inhibitors to control weedy rice. Weedy rice plants have differential tolerance to imazethapyr, perhaps due to ALS polymorphisms. We aimed to assess ALS polymorphisms in weedy rice accessions from Arkansas, USA, with differential tolerance to imazethapyr in seedling growth bioassays. Six base changes were identified in the ALS of 14 weedy rice accessions. Three of these nucleotide changes resulted in amino acid substitutions — Pro⁹³Thr, Glu⁶³⁰Asp, and Val⁶⁶⁹Met — in four accessions: Ark-4, Ark-9, Poi-1 and Poi-6. The GR₅₀ values and inhibition of root and shoot growth (%) of these accessions differed. The Glu⁶³⁰Asp substitution occurs in the herbicide binding domain B and Val⁶⁶⁹Met occurs at the C-terminal tail where the co-factor binds. Variability in weedy rice ALS exists, but polymorphism patterns did not relate to tolerance levels. The observed mutations presented the possibility that sustained selection pressure will eventually lead to selection of a herbicide-tolerant individual that will be the progenitor of a resistant population. Concomitantly, pollen-mediated gene flow from Clearfield™ rice to weedy rice will lead to the evolution of ALS-resistant weedy rice populations.     

Development of simple and co-dominant PCR markers to genotype puroindoline a and b alleles for grain hardness in bread wheat (Triticum aestivum L.)

Grain hardness is one of the most important quality characteristics of cultivated bread wheat (Triticum aestivum L.). A large deletion in the puroindoline a (Pina) gene or single nucleotide polymorphisms (SNPs) in the puroindoline b (Pinb) gene results in hard grain texture. So far, nine Pina alleles (Pina-D1a–Pina-D1b, Pina-D1k–Pina-D1q) and seventeen Pinb alleles (Pinb-D1a–Pinb-D1g, Pinb-D1p–Pinb-D1ab) have been identified in bread wheat. The major Pina and Pinb alleles identified in hard wheat cultivars are Pina-D1b, Pinb-D1b, Pinb-D1c and Pinb-D1d. In this study, a three-primer PCR system was employed to develop nine co-dominant STS markers for genotyping Pina-D1a and Pina-D1b, whereas temperature-switch (TS) PCR was used to develop six co-dominant SNP markers for genotyping the Pinb-D1a, Pinb-D1b, Pinb-D1c and Pinb-D1d alleles. These STS and TS-PCR markers were used to verify the grain hardness genotype of 100 wheat cultivars. The reliability and genotyping accuracy of TS-PCR markers were confirmed through sequencing of PCR products and a comparison with previously published results. Therefore, STS and TS-PCR markers offer a simple, cost-effective and reliable method for high-throughput genotyping Pina and Pinb alleles to select grain hardness in wheat quality breeding programs and for wheat market classification.