Molecular and phylogenetic characterization of the homoeologous EPSP Synthase genes of allohexaploid wheat, Triticum aestivum (L.).

BACKGROUND: 5-Enolpyruvylshikimate-3-phosphate synthase (EPSPS) is the sixth and penultimate enzyme in the shikimate biosynthesis pathway, and is the target of the herbicide glyphosate. The EPSPS genes of allohexaploid wheat (Triticum aestivum, AABBDD) have not been well characterized. Herein, the three homoeologous copies of the allohexaploid wheat EPSPS gene were cloned and characterized. METHODS: Genomic and coding DNA sequences of EPSPS from the three related genomes of allohexaploid wheat were isolated using PCR and inverse PCR approaches from soft white spring "Louise'. Development of genome-specific primers allowed the mapping and expression analysis of TaEPSPS-7A1, TaEPSPS-7D1, and TaEPSPS-4A1 on chromosomes 7A, 7D, and 4A, respectively. Sequence alignments of cDNA sequences from wheat and wheat relatives served as a basis for phylogenetic analysis. RESULTS: The three genomic copies of wheat EPSPS differed by insertion/deletion and single nucleotide polymorphisms (SNPs), largely in intron sequences. RT-PCR analysis and cDNA cloning revealed that EPSPS is expressed from all three genomic copies. However, TaEPSPS-4A1 is expressed at much lower levels than TaEPSPS-7A1 and TaEPSPS-7D1 in wheat seedlings. Phylogenetic analysis of 1190-bp cDNA clones from wheat and wheat relatives revealed that: 1) TaEPSPS-7A1 is most similar to EPSPS from the tetraploid AB genome donor, T. turgidum (99.7 % identity); 2) TaEPSPS-7D1 most resembles EPSPS from the diploid D genome donor, Aegilops tauschii (100 % identity); and 3) TaEPSPS-4A1 resembles EPSPS from the diploid B genome relative, Ae. speltoides (97.7 % identity). Thus, EPSPS sequences in allohexaploid wheat are preserved from the most two recent ancestors. The wheat EPSPS genes are more closely related to Lolium multiflorum and Brachypodium distachyon than to Oryza sativa (rice). CONCLUSIONS: The three related EPSPS homoeologues of wheat exhibited conservation of the exon/intron structure and of coding region sequence, but contained significant sequence variation within intron regions. The genome-specific primers developed will enable future characterization of natural and induced variation in EPSPS sequence and expression. This can be useful in investigating new causes of glyphosate herbicide resistance.

Increased ABA sensitivity results in higher seed dormancy in soft white spring wheat cultivar 'Zak'.

As a strategy to increase the seed dormancy of soft white wheat, mutants with increased sensitivity to the plant hormone abscisic acid (ABA) were identified in mutagenized grain of soft white spring wheat "Zak". Lack of seed dormancy is correlated with increased susceptibility to preharvest sprouting in wheat, especially those cultivars with white kernels. ABA induces seed dormancy during embryo maturation and inhibits the germination of mature grain. Three mutant lines called Zak ERA8, Zak ERA19A, and Zak ERA19B (Zak ENHANCED RESPONSE to ABA) were recovered based on failure to germinate on 5 µM ABA. All three mutants resulted in increased ABA sensitivity over a wide range of concentrations such that a phenotype can be detected at very low ABA concentrations. Wheat loses sensitivity to ABA inhibition of germination with extended periods of dry after-ripening. All three mutants recovered required more time to after-ripen sufficiently to germinate in the absence of ABA and to lose sensitivity to 5 µM ABA. However, an increase in ABA sensitivity could be detected after as long as 3 years of after-ripening using high ABA concentrations. The Zak ERA8 line showed the strongest phenotype and segregated as a single semi-dominant mutation. This mutation resulted in no obvious decrease in yield and is a good candidate gene for breeding preharvest sprouting tolerance.

Scarlet-Rz1, an EMS-generated hexaploid wheat with tolerance to the soilborne necrotrophic pathogens Rhizoctonia solani AG-8 and R. oryzae.

The necrotrophic root pathogens Rhizoctonia solani AG-8 and R. oryzae cause Rhizoctonia root rot and damping-off, yield-limiting diseases that pose barriers to the adoption of conservation tillage in wheat production systems. Existing control practices are only partially effective, and natural genetic resistance to Rhizoctonia has not been identified in wheat or its close relatives. We report the first genetic resistance/tolerance to R. solani AG-8 and R. oryzae in wheat (Triticum aestivum L. em Thell) germplasm 'Scarlet-Rz1'. Scarlet-Rz1 was derived from the allohexaploid spring wheat cultivar Scarlet using EMS mutagenesis. Tolerant seedlings displayed substantial root and shoot growth after 14 days in the presence of 100-400 propagules per gram soil of R. solani AG-8 and R. oryzae in greenhouse assays. BC(2)F(4) individuals of Scarlet-Rz1 showed a high and consistent degree of tolerance. Seedling tolerance was transmissible and appeared to be dominant or co-dominant. Scarlet-Rz1 is a promising genetic resource for developing Rhizoctonia-tolerant wheat cultivars because the tolerance trait immediately can be deployed into wheat breeding germplasm through cross-hybridization, thereby avoiding difficulties with transfer from secondary or tertiary relatives as well as constraints associated with genetically modified plants. Our findings also demonstrate the utility of chemical mutagenesis for generating tolerance to necrotrophic pathogens in allohexaploid wheat.

Influence of glyphosate, crop volunteer and root pathogens on glyphosate-resistant wheat under controlled environmental conditions.

BACKGROUND: The herbicide glyphosate has a synergistic effect on root disease because of increased susceptibility from reduced plant defenses resulting from the blockage of the shikimic acid pathway. Could glyphosate-resistant (GR) wheat cultivars and glyphosate application in-crop increase the risk of damage from soil-borne pathogens? Growth chamber experiments were conducted with two GR wheat lines and their corresponding glyphosate-sensitive (GS) parents and four pathogens (Rhizoctonia solani Kühn R. oryzae Ryker & Gooch, Gaeumannomyces graminis (Sacc.) v. Arx & J. Olivier var. tritici J. Walker and Pythium ultimum Trow). Treatments consisted of different herbicide timings and planting of crop volunteer to mimic management practices in the field. RESULTS: GR cultivars were not inherently more susceptible to root pathogens than GS cultivars, and application of glyphosate did not increase root disease. When crop volunteer was grown in close proximity to GR cultivars, the timing of glyphosate application had a profound effect. In general, the longer the crop volunteer was left before killing with glyphosate, the greater was the competitive effect on the planted crop. Both R. solani and G. graminis var. tritici reduced plant height, number of tillers and root length of the GR cultivars in the presence of crop volunteer with glyphosate application. CONCLUSION: To minimize the damaging effects of these pathogens, producers should apply glyphosate at least 2-3 weeks before planting GR wheat, as currently advised for GS cereals.

Economically Optimal Wheat Yield, Protein and Nitrogen Use Component Responses to Varying N Supply and Genotype.

Improvements in market value of hard red spring wheat (HRS, Triticum aestivum L.) are linked to breeding efforts to increase grain protein concentration (GPC). Numerous studies have been conducted on the identification, isolation of a chromosome region (Gpc-B1) of Wild emmer wheat (Triticum turgidum spp. dicoccoides) and its introgression into commercial hard wheat to GPC. Yet there has been limited research published on the comparative responsiveness of these altered lines and their parents to varied N supply. There is increased awareness that wheat genetic improvements must be assessed over a range of environmental and agronomic management conditions to assess stability. We report herein on economically optimal yield, protein and nitrogen use efficiency (NUE) component responses of two Pacific Northwestern USA cultivars, Tara and Scarlet compared to backcrossed derived near isolines with or without the Gpc-B1 allele. A field experiment with 5 N rates as whole plots and 8 genotypes as subplots was conducted over two years under semi-arid, dryland conditions. One goal was to evaluate the efficacy of the Gpc-B1 allele under a range of low to high N supply. Across all genotypes, grain yield responses to N supply followed the classic Mitscherlich response model, whereas GPC followed inverse quadratic or linear responses. The Gpc-B1 introgression had no major impact on grain protein, but grain N and total above ground crop N yields demonstrated quadratic responses to total N supply. Generally, higher maximum grain yields and steeper rise to the maxima (Mitscherlich c values) were obtained in the first site-year. Tara required less N supply to achieve GPC goals than Scarlet in both site-years. Genotypes with Gpc-B1 produced comparable or slightly lower Mitscherlich A values than unmodified genotypes, but displayed similar Mitscherlich c values. Target GPC goals were not achieved at economic optimal yields based on set wheat pricing. Economic optimization of N inputs to achieve protein goals showed positive revenue from additional N inputs for most genotypes. While N uptake efficiency did not drop below 0.40, N fertilizer-induced increases in grain N harvest correlated well with unused post-harvest soil N that is potentially susceptible to environmental loss.

Proso Millet (Panicum miliaceum L.) and Its Potential for Cultivation in the Pacific Northwest, U.S.: A Review.

Proso millet (Panicum miliaceum L.) is a warm season grass with a growing season of 60-100 days. It is a highly nutritious cereal grain used for human consumption, bird seed, and/or ethanol production. Unique characteristics, such as drought and heat tolerance, make proso millet a promising alternative cash crop for the Pacific Northwest (PNW) region of the United States. Development of proso millet varieties adapted to dryland farming regions of the PNW could give growers a much-needed option for diversifying their predominantly wheat-based cropping systems. In this review, the agronomic characteristics of proso millet are discussed, with emphasis on growth habits and environmental requirements, place in prevailing crop rotations in the PNW, and nutritional and health benefits. The genetics of proso millet and the genomic resources available for breeding adapted varieties are also discussed. Last, challenges and opportunities of proso millet cultivation in the PNW are explored, including the potential for entering novel and regional markets.

Economic impact of Hessian fly (Diptera: Cecidomyiidae) on spring wheat in Oregon and additive yield losses with Fusarium crown rot and lesion nematode.

Damage caused by Hessian fly, Mayetiola destructor (Say), was quantified in spring wheat, Triticum aestivum L., trials near Pendleton and Moro, OR, during 2001 and 2002. Five field experiments were established to examine genetic resistance to Fusarium crown rot, Fusarium pseudograminearum (O'Donnell & Aoki), and economic damage by lesion nematode, Pratylenchus neglectus (Rensch, 1924) (Filipjev Schuurmanns & Stekhoven, 1941) and Pratylenchus thornei (Sher & Allen, 1941). Hessian fly became the dominant factor affecting grain yield in four experiments. Genotypes carrying the H3-resistance gene had grain yields 66 and 68% higher than susceptible genotypes in cultivar trials during 2001 and 2002, respectively. Yield reductions were detected when Hessian fly infestation rates exceeded 50% plants during 2001 and 15% plants (8% tillers) during 2002. In two trials during 2001, in-furrow application of aldicarb (Temik) at planting improved yields of four Hessian fly-susceptible cultivars by 72 and 144% (up to 1,959 kg/ha) and yields of one Hessian fly-resistant cultivar by 2 and 3%. Resistant cultivars and aldicarb improved grain quality as much as two market grades during 2001. The value of increased grain production with Hessian fly-resistant cultivars in four field experiments ranged from dollar 112 to dollar 252/ha, excluding price incentives for improved market quality. Yield reduction due to combined damage from Hessian fly and either Fusarium crown rot or lesion nematode was additive. This report seems to be the first quantitative yield loss estimate for Hessian fly in spring wheat in the semiarid environment of the inland Pacific Northwest.

Registration of Zak ERA8 Soft White Spring Wheat Germplasm with Enhanced Response to ABA and Increased Seed Dormancy.

Zak ERA8 (ENHANCED RESPONSE to ABA8) (Reg. No. GP-966, PI 669443) is a unique line derived from soft white spring wheat (Triticum aestivum L.) cultivar Zak that has increased seed dormancy but after-ripens within 10 to 16 wk. The goal in developing this germplasm was to use increased seed dormancy to improve tolerance to preharvest sprouting, a problem that can cause severe economic losses. This germplasm was developed by USDA-ARS, Pullman, WA, in collaboration with Washington State University. Zak ERA8was tested under experimental number 60.1.27.10. The ERA8mutation was generated by chemical mutagenesis followed by selection for the inability to germinate on abscisic acid (ABA) concentrations too low to inhibit wild-type Zak seed germination. The semidominant Zak ERA8 line has been backcrossed twice to wild-type Zak. Following the first backcross, Zak ERA8 showed similar morphological and grain quality traits to the original Zak cultivar.

Glyphosate inhibits rust diseases in glyphosate-resistant wheat and soybean.

Glyphosate is a broad-spectrum herbicide used for the control of weeds in glyphosate-resistant crops. Glyphosate inhibits 5-enolpyruvyl shikimate 3-phosphate synthase, a key enzyme in the synthesis of aromatic amino acids in plants, fungi, and bacteria. Studies with glyphosate-resistant wheat have shown that glyphosate provided both preventive and curative activities against Puccinia striiformis f. sp. tritici and Puccinia triticina, which cause stripe and leaf rusts, respectively, in wheat. Growth-chamber studies demonstrated wheat rust control at multiple plant growth stages with a glyphosate spray dose typically recommended for weed control. Rust control was absent in formulation controls without glyphosate, dependent on systemic glyphosate concentrations in leaf tissues, and not mediated through induction of four common systemic acquired resistance genes. A field test with endemic stripe rust inoculum confirmed the activities of glyphosate pre- and postinfestation. Preliminary greenhouse studies also demonstrated that application of glyphosate in glyphosate-resistant soybeans suppressed Asian soybean rust, caused by Phakopsora pachyrhizi.