Transcriptional regulation mechanism of wheat varieties with different nitrogen use efficiencies in response to nitrogen deficiency stress.

BACKGROUND: As one of the microelements, nitrogen play essential roles in cereal production. Although the use of chemical fertilizers has significantly improved the yield of wheat, it has also caused increasingly adverse environmental pollution. Revealing the molecular mechanism manipulating wheat nitrogen use efficiency (NUE), and cultivating wheat germplasms with high nitrogen use efficiency has become important goals for wheat researchers. In this study, we investigated the physiological and transcriptional differences of three wheat cultivars with different NUE under low nitrogen stress. RESULTS: The results showed that, under low nitrogen conditions, the activities of nitrogen metabolism-related enzymes (GS, NR, GDH), antioxidant enzymes (SOD, POD, CAT) and soluble protein contents of ZM366 (high NUE cultivar) were higher than those of JD8 (low NUE cultivar). The hybrid cultivar of ZM366 and JD8 showed mid-parent or over-parent heterosis. Transcriptome analysis revealed that 'alanine, aspartate and glutamate metabolism', 'terpenoid backbone biosynthesis' and 'vitamin B6 metabolism' pathways play key roles in nitrogen use efficiency in wheat. The significant enhancement of the 'Calvin cycle' and 'photorespiration' in ZM366 contributed to its higher level of carbon metabolism under low nitrogen stress, which is an important attribute differs from the other two varieties. In addition, the activation of ABA signal transduction and biosynthesis pathways also helps to maintain NUE under low- nitrogen conditions. Moreover, bHLH transcription factors were also found to play a positive role in wheat NUE. CONCLUSIONS: In conclusion, these results enriched our knowledge of the mechanism of wheat NUE, and provided a theoretical basis for improving wheat NUE and breeding new cultivars.

Over-Expressing TaSPA-B Reduces Prolamin and Starch Accumulation in Wheat (Triticum aestivum L.) Grains.

Starch and prolamin composition and content are important indexes for determining the processing and nutritional quality of wheat (Triticum aestivum L.) grains. Several transcription factors (TFs) regulate gene expression during starch and protein biosynthesis in wheat. Storage protein activator (TaSPA), a member of the basic leucine zipper (bZIP) family, has been reported to activate glutenin genes and is correlated to starch synthesis related genes. In this study, we generated TaSPA-B overexpressing (OE) transgenic wheat lines. Compared with wild-type (WT) plants, the starch content was slightly reduced and starch granules exhibited a more polarized distribution in the TaSPA-B OE lines. Moreover, glutenin and ω- gliadin contents were significantly reduced, with lower expression levels of related genes (e.g., By15, Dx2, and ω-1,2 gliadin gene). RNA-seq analysis identified 2023 differentially expressed genes (DEGs). The low expression of some DEGs (e.g., SUSase, ADPase, Pho1, Waxy, SBE, SSI, and SS II a) might explain the reduction of starch contents. Some TFs involved in glutenin and starch synthesis might be regulated by TaSPA-B, for example, TaPBF was reduced in TaSPA-B OE-3 lines. In addition, dual-luciferase reporter assay indicated that both TaSPA-B and TaPBF could transactivate the promoter of ω-1,2 gliadin gene. These results suggest that TaSPA-B regulates a complex gene network and plays an important role in starch and protein biosynthesis in wheat.

Expression of Pinellia pedatisecta Lectin Gene in Transgenic Wheat Enhances Resistance to Wheat Aphids.

Wheat aphids are major pests during the seed filling stage of wheat. Plant lectins are toxic to sap-sucking pests such as wheat aphids. In this study, Pinellia pedatisecta agglutinin (ppa), a gene encoding mannose binding lectin, was cloned, and it shared 92.69% nucleotide similarity and 94% amino acid similarity with Pinellia ternata agglutinin (pta). The ppa gene, driven by the constitutive and phloem-specific ribulose bisphosphate carboxylase small subunit gene (rbcs) promoter in pBAC-rbcs-ppa expression vector, was transferred into the wheat cultivar Baofeng104 (BF104) by particle bombardment transformation. Fifty-four T0 transgenic plants were generated. The inheritance and expression of the ppa gene were confirmed by PCR and RT-PCR analysis respectively, and seven homozygous transgenic lines were obtained. An aphid bioassay on detached leaf segments revealed that seven ppa transgenic wheat lines had lower aphid growth rates and higher inhibition rates than BF104. Furthermore, two-year aphid bioassays in isolated fields showed that aphid numbers per tiller of transgenic lines were significantly decreased, compared with wild type BF104. Therefore, ppa could be a strong biotechnological candidate to produce aphid-resistant wheat.

Silencing of a lipase maturation factor 2-like gene by wheat-mediated RNAi reduces the survivability and reproductive capacity of the grain aphid, Sitobion avenae.

Lipase maturation factor (LMF) family proteins are required for the maturation and transport of active lipoprotein lipases. However, the specific roles of LMF2 remain unknown. In this study, a grain aphid lmf2-like gene fragment was cloned and was highly similar in sequence to a homologous gene in the pea aphid, Acyrthosiphon pisum. An RNAi vector was constructed with this fragment and used for wheat transformation. The expression of the lmf2-like gene in aphid, as well as the growth and reproduction of the aphids, was analyzed after feeding on the transgenic wheat. There were no significant differences in the expression of the lmf2-like gene over development. The expression of the lmf2-like gene was significantly reduced by 27.6% on the fifth day, and 57.6% on the 10th day after feeding. The total number of aphids produced on the transgenic plants was less than the number produced on control plants, and the difference became significant or after 2 weeks. The molting numbers were also reduced in the aphids reared on the transgenic plants. Our findings indicate that lmf2-like genes may have potential as a target gene for the control of grain aphids and show that feeding aphids with wheat expressing lmf2-like RNAi resulted in significant reductions in survival and reproduction.

Detection of seed purity of hybrid wheat using reflectance and transmittance hyperspectral imaging technology.

Chemical hybridization and genic male sterility systems are two main methods of hybrid wheat production; however, complete sterility of female wheat plants cannot be guaranteed owing to the influence of the growth stage and weather. Consequently, hybrid wheat seeds are inevitably mixed with few parent seeds, especially female seeds. Therefore, seed purity is a key factor in the popularization of hybrid wheat. However, traditional seed purity detection and variety identification methods are time-consuming, laborious, and destructive. Therefore, to establish a non-destructive classification method for hybrid and female parent seeds, three hybrid wheat varieties (Jingmai 9, Jingmai 11, and Jingmai 183) and their parent seeds were sampled. The transmittance and reflectance spectra of all seeds were collected via hyperspectral imaging technology, and a classification model was established using partial least squares-discriminant analysis (PLS-DA) combined with various preprocessing methods. The transmittance spectrum significantly improved the classification of hybrids and female parents compared to that obtained using reflectance spectrum. Specifically, using transmittance spectrum combined with a characteristic wavelength-screening algorithm, the Detrend-CARS-PLS-DA model was established, and the accuracy rates in the testing sets of Jingmai 9, Jingmai 11, and Jingmai 183 were 95.69%, 98.25%, and 97.25%, respectively. In conclusion, transmittance hyperspectral imaging combined with a machine learning algorithm can effectively distinguish female parent seeds from hybrid seeds. These results provide a reference for rapid seed purity detection in the hybrid production process. Owing to the non-destructive and rapid nature of hyperspectral imaging, the detection of hybrid wheat seed purity can be improved by online sorting in the future.

Plant-mediated gene silencing of an essential olfactory-related Gqα gene enhances resistance to grain aphid in common wheat in greenhouse and field.

BACKGROUND: Grain aphid (Sitobion avenae F.) is a dominant pest that limits cereal crop production around the globe. Gq proteins have important roles in signal transduction in insect olfaction. Plant-mediated RNA interference (RNAi) has been widely studied in insect control, but its application for the control wheat aphid in the field requires further study. Here, we used double-stranded (ds)RNA feeding to verify the potential of selected Gqα fragments for host-mediated RNAi, and then evaluated the effect of RNAi on aphid olfaction in transgenic wheat in the greenhouse and field. RESULTS: Gqα gene was expressed in the aphid life cycle, and a 540 bp fragment shared 98.1% similarity with the reported sequence. dsGqα feeding reduced the expression of Gqα, and both reproduction and molting in the grain aphid. Feeding transgenic lines in the greenhouse downregulated expression of aphid Gqα, and significantly reduced reproduction and molting numbers. Furthermore, our field results indicate that transgenic lines have lower aphid numbers and higher 1000-grain weight than an unsprayed wild-type control. CONCLUSION: Plant-mediated silencing of an essential olfactory-related Gqα gene could enhance resistance to grain aphid in common wheat in both the greenhouse and the field. © 2018 Society of Chemical Industry.