Functional analysis of polyphenol oxidase 1 gene in common wheat.

Polyphenol oxidase (PPO) activity is a major cause of the undesirable brown color of wheat-based products. Ppo1, a major gene for PPO activity, was cloned based on sequence homology in previous studies; however, its function and regulation mechanism remain unclear. In this study, the function and genetic regulation of Ppo1 were analyzed using RNA interference (RNAi) and Targeting Induced Local Lesions IN Genomes (TILLING) technology, and superior mutants were identified. Compared with the control, the level of Ppo1 transcript in RNAi transgenic lines was drastically decreased by 15.5%-60.9% during grain development, and PPO activity was significantly reduced by 12.9%-20.4%, confirming the role of Ppo1 in PPO activity. Thirty-two Ppo1 mutants were identified in the ethyl methanesulfonate (EMS)-mutagenized population, including eight missense mutations, 16 synonymous mutations, and eight intron mutations. The expression of Ppo1 was reduced significantly by 6.7%-37.1% and 10.1%-54.4% in mutants M092141 (G311S) and M091098 (G299R), respectively, in which PPO activity was decreased by 29.7% and 28.8%, respectively, indicating that mutation sites of two mutants have important effects on PPO1 function. Sequence and structure analysis revealed that the two sites were highly conserved among 74 plant species, where the frequency of glycine was 94.6% and 100%, respectively, and adjacent to the entrance of the hydrophobic pocket of the active site. The M092141 and M091098 mutants can be used as important germplasms to develop wheat cultivars with low grain PPO activity. This study provided important insights into the molecular mechanism of Ppo1 and the genetic improvement of wheat PPO activity.

Slow stripe rusting in Chinese wheat Jimai 44 conferred by Yr29 in combination with a major QTL on chromosome arm 6AL.

KEY MESSAGE: YrJ44, a more effective slow rusting gene than Yr29, was localized to a 3.5-cM interval between AQP markers AX-109373479 and AX-109563479 on chromosome 6AL. "Slow rusting" (SR) is a type of adult plant resistance (APR) that can provide non-specific durable resistance to stripe rust in wheat. Chinese elite wheat cultivar Jimai 44 (JM44) has maintained SR to stripe rust in China since its release despite exposure to a changing and variable pathogen population. An F2:6 population comprising 295 recombinant inbred lines (RILs) derived from a cross between JM44 and susceptible cultivar Jimai 229 (JM229) was used in genetic analysis of the SR. The RILs and parental lines were evaluated for stripe rust response in five field environments and genotyped using the Affymetrix Wheat55K SNP array and 13 allele-specific quantitative PCR-based (AQP) markers. Two stable QTL on chromosome arms 1BL and 6AL were identified by inclusive composite interval mapping. The 1BL QTL was probably the pleiotropic gene Lr46/Yr29/Sr58. QYr.nwafu-6AL (hereafter named YrJ44), mapped in a 3.5-cM interval between AQP markers AX-109373479 and AX-109563479, was more effective than Yr29 in reducing disease severity and relative area under the disease progress curve (rAUDPC). RILs harboring both YrJ44 and Yr29 displayed levels of SR equal to the resistant parent JM44. The AQP markers linked with YrJ44 were polymorphic and significantly correlated with stripe rust resistance in a panel of 1,019 wheat cultivars and breeding lines. These results suggested that adequate SR resistance can be obtained by combining YrJ44 and Yr29 and the AQP markers can be used in breeding for durable stripe rust resistance.

Moderate addition of B-type starch granules improves the rheological properties of wheat dough.

The quality traits of wheat grain ultimately determine the performance of wheat flour and dough, which is crucial to end-products. However, to combine high yield and good grain quality has been a great challenge in wheat breeding. In this study, the different sized A- and B-type starch granules were fractioned to investigate their effects on the physicochemical properties of wheat flour and rheological properties of wheat dough using three substitution levels (5, 10, and 15%). Results showed that 5% B-type starch granules addition increased the percentage of SDS-unextractable polymeric protein, optimized the dough network, and increased the bond water content, and thus improved the dough rheological properties. The addition of A-type starch granules or excessive B-type starch granules diluted and destroyed the structure of gluten, and reduced the dough strength. Therefore, a possible strategy for combining wheat quality and yield was proposed, that is, replacing protein content with B-type starch granules at a proper level, which has profound implications for wheat breeders to look at and address trade-offs between the quality and yield of wheat in future.

Increasing the Grain Yield and Grain Protein Content of Common Wheat (Triticum aestivum) by Introducing Missense Mutations in the Q Gene.

Grain yield (GY) and grain protein content (GPC) are important traits for wheat breeding and production; however, they are usually negatively correlated. The Q gene is the most important domestication gene in cultivated wheat because it influences many traits, including GY and GPC. Allelic variations in the Q gene may positively affect both GY and GPC. Accordingly, we characterized two new Q alleles (Qs1 and Qc1-N8) obtained through ethyl methanesulfonate-induced mutagenesis. Compared with the wild-type Q allele, Qs1 contains a missense mutation in the sequence encoding the first AP2 domain, whereas Qc1-N8 has two missense mutations: one in the sequence encoding the second AP2 domain and the other in the microRNA172-binding site. The Qs1 allele did not significantly affect GPC or other processing quality parameters, but it adversely affected GY by decreasing the thousand kernel weight and grain number per spike. In contrast, Qc1-N8 positively affected GPC and GY by increasing the thousand kernel weight and grain number per spike. Thus, we generated novel germplasm relevant for wheat breeding. A specific molecular marker was developed to facilitate the use of the Qc1-N8 allele in breeding. Furthermore, our findings provide useful new information for enhancing cereal crops via non-transgenic approaches.

Amino acid transporter gene TaATLa1 from Triticum aestivum L. improves growth under nitrogen sufficiency and is down regulated under nitrogen deficiency.

MAIN CONCLUSION: TaATLa1 was identified to respond to nitrogen deprivation through transcriptome analysis of wheat seedlings. TaATLa1 specifically transports Gln, Glu, and Asp, and affects the biomass of Arabidopsis and wheat. Nitrogen is an essential macronutrient and plays a crucial role in wheat production. Amino acids, the major form of organic nitrogen, are remobilized by amino acid transporters (AATs) in plants. AATs are commonly described as central components of essential developmental processes and yield formation via taking up and transporting amino acids in plants. However, few studies have reported the detailed biochemical properties and biological functions of these AATs in wheat. In this study, key genes encoding AATs were screened from transcriptome analysis of wheat seedlings treated with normal nitrogen (NN) and nitrogen deprivation (ND). Among them, 21 AATs were down-regulated and eight AATs were up-regulated under ND treatment. Among the homoeologs, TaATLa1.1-3A, TaATLa1.1-3B, and TaATLa1.1-3D (TaATLa1.1-3A, -3B, and -3D), belonging to amino acid transporter-like a (ATLa) subfamily, were significantly down-regulated in response to ND in wheat, and accordingly were selected for functional analyses. The results demonstrated that TaATLa1.1-3A, -3B, and -3D effectively transported glutamine (Gln), glutamate (Glu), and aspartate (Asp) in yeast. Overexpression of TaAILa1.1-3A, -3B, and -3D in Arabidopsis thaliana L. significantly increased amino acid content in leaves, storage protein content in seeds and the plant biomass under NN. Knockdown of TaATLa1.1-3A, -3B, and -3D in wheat seedlings resulted in a significant block of amino acid remobilization and growth inhibition. Taken together, TaATLa1.1-3A, -3B, and -3D contribute substantially to Arabidopsis and wheat growth. We propose that TaATLa1.1-3A, -3B, and -3D may participate in the source-sink translocation of amino acid, and they may have profound implications for wheat yield improvement.

Mitogen-activated protein kinase TaMPK3 suppresses ABA response by destabilising TaPYL4 receptor in wheat.

Abscisic acid (ABA) receptors are considered as the targeted manipulation of ABA sensitivity and water productivity in plants. Regulation of their stability or activity will directly affect ABA signalling. Mitogen-activated protein kinase (MAPK) cascades link multiple environmental and plant developmental cues. However, the molecular mechanism of ABA signalling and MAPK cascade interaction remains largely elusive. TaMPK3 overexpression decreases drought tolerance and wheat sensitivity to ABA, significantly weakening ABA's inhibitory effects on growth. Under drought stress, overexpression lines show lower survival rates, shoot fresh weight and proline content, but higher malondialdehyde levels at seedling stage, as well as decreased grain width and 1000 grain weight in both glasshouse and field conditions at the adult stage. TaMPK3-RNAi increases drought tolerance. TaMPK3 interaction with TaPYL4 leads to decreased TaPYL4 levels by promoting its ubiquitin-mediated degradation, whereas ABA treatment diminishes TaMPK3-TaPYL interactions. In addition, the expression of ABA signalling proteins is impaired in TaMPK3-overexpressing wheat plants under ABA treatment. The MPK3-PYL interaction module was found to be conserved across monocots and dicots. Our results suggest that the MPK3-PYL module could serve as a negative regulatory mechanism for balancing appropriate drought stress response with normal plant growth signalling in wheat.

Effects of Quinoa Flour on Wheat Dough Quality, Baking Quality, and in vitro Starch Digestibility of the Crispy Biscuits.

Quinoa is a pseudo-cereal which has excellent nutritional and functional properties due to its high content of nutrients, such as polyphenols and flavonoids, and therefore quinoa serves as an excellent supplement to make healthy and functional foods. The present study was aimed to evaluate the quality characteristics of wheat doughs and crispy biscuits supplemented with different amount of quinoa flour. The results showed that when more wheat flour was substituted by quinoa flour, proportion of unextractable polymeric protein to the total polymeric protein (UPP%) of the reconstituted doughs decreased and the gluten network structure was destroyed at a certain substitution level. The content of B-type starch and the gelatinization temperature of the reconstituted flours increased. The storage modulus, loss modulus, development time, and stability time of the dough increased as well. Moreover, hardness and toughness of the formulated crispy biscuits significantly decreased. Analyses suggested that starch digestibility was reduced and resistant starch content increased significantly. Taken together, quinoa flour improved dough rheological properties, enhanced the textural properties, and increased resistant starch content in crispy biscuits, thus adding to high nutritional value.

Effect of Highland Barley on Rheological Properties, Textural Properties and Starch Digestibility of Chinese Steamed Bread.

Highland barley has a different composition and structure to other crops. It has higher contents of total polyphenol (TPC), total flavonoid (TFC) and ß-glucan, which can be supplemented to improve the nutrition of wheat-flour-based food. In this study, the flours of three different grain-colored highland barley varieties Beiqing 6 (BQ), Dulihuang (DLH), and Heilaoya (HLY), were added to Jimai60 (JM, a wheat variety with medium gluten) wheat flour at different substitution levels to investigate their effects on the unextractable polymeric protein (UPP) content, micro-structure, rheological properties and mixing properties of dough, and the color, texture, flavor, and in vitro digestion of Chinese steam bread (CSB). The results showed that the moderate substitution of highland barley (20%) increased the UPP%, optimized the micro-structure of gluten, and improved its rheological properties by increasing dough viscoelasticity. The CSBs made from the composite flours exhibited a similar specific volume, cohesiveness, springiness and resilience to wheat CSB, while the firmness of composite CSBs (particularly JM-HLY-20) was delayed during storage. Importantly, the addition of highland barley increased the contents of TPC, TFC and ß-glucan, but decreased the in vitro starch digestibility of CSBs. A sensory evaluation showed that JM-HLY CSB was the most preferable. Taken together, highland barley can be used as a fine supplement to food products, with health-promoting properties.

Interaction of B-type starch with gluten skeleton improves wheat dough mixing properties by stabilizing gluten micro-structure.

Some recent studies have revealed individual and the combined interactions of gluten and starch affecting dough mixing properties. However, the combined influence of high-molecular-weight glutenin subunits (HMW-GS) and starch on dough mixing and rheological properties requires elucidation. Thus four recombinant inbred lines, SS 1, SS 2, ZZ 1 and ZZ 2, were selected based on their HMW-GSs compositions. Compared to ZZ 1 and ZZ 2, both SS 1 and SS 2 carried superior HMW-GS alleles, and exhibited extended dough development and stability time, indicating their significant dough mixing characteristics. The gluten skeleton of the wheat lines SS 2 and ZZ 2 with higher B-type starch proportions exhibited fewer breakages along with the rise of dough temperature during mixing. Higher content of B-type starch strengthens interaction between starch and gluten skeleton at the dough heating stage, suggesting a specific range of B-type starch proportion can improve dough mixing characteristics.

Effects of the different waxy proteins on starch biosynthesis, starch physicochemical properties and Chinese noodle quality in wheat.

Noodles are an important food in Asia. Wheat starch is the most important component in Chinese noodles. Loss of the waxy genes leads to lower activity of starch synthesis enzymes and decreased amylose content that further affects starch properties and noodle quality. To study the effects of different waxy (Wx) protein subunits on starch biosynthesis and processing quality, the high-yielding wheat cultivar Jimai 22 was treated with the mutagen ethyl methane sulfonate (EMS) to produce a population of Wx lines and chosen 7 Wx protein combinations. The amylose content increased but swelling power decreased as the number of Wx proteins increased. Both GBSS activity and gene expression were the lowest for the waxy mutant, followed by the mutants with 1 Wx protein. The combinations of these mutant alleles lead to reductions in both RNA expression and protein levels. Noodles made from materials with 2 Wx protein subunits had the highest score, which agreed with peak viscosity. The influence of the Wx-B1 protein on amylose synthesis and noodle quality was the highest, whereas the influence of Wx-A1 protein was the lowest. Mutants with lower amylose content caused by the absence of 1 subunit, especially the Wx-B1 subunit, had superior noodle quality. Additionally, the identified mutant lines can be used as intermediate materials to improve wheat quality. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01292-x.

Effect of Hulless Barley Flours on Dough Rheological Properties, Baking Quality, and Starch Digestibility of Wheat Bread.

Hulless barley (Hordeum vulgare L.), also known as highland barley, contains nutritional compounds, such as ß-glucan and polyphenol, which can be added to wheat flour to improve the dough nutritional quality. In this study, different formulated dough samples were obtained by individually adding four hulless barley flours into flour of a wheat variety (Jimai 44, designated as JM) which has very strong gluten. The effects of hulless barley supplementation on gluten structure, dough rheological properties, bread-making properties, and starch digestibility were assessed. The results showed that compared with JM dough, substitution of hulless barley flour to wheat flour at levels ranging from 10 to 40% negatively affected gluten micro-structure and dough mixing behavior, because the cross-links of gluten network were partially broken and the dough development time and stability time were shortened. For the hulless barley-supplemented bread, specific volume was significantly (P < 0.05) increased while springiness was not greatly changed. Furthermore, the hydrolysed starch rate in hulless barley-supplemented bread was decreased, compared with that in JM bread. Importantly, the contents of ß-glucan, polyphenols and flavonoids in hulless barley-supplemented bread were 132.61-160.87%, 5.71-48.57%, and 25-293.75% higher than those in JM bread, respectively. Taken together, the hulless barley-supplemented bread has been fortified with enhanced nutritional components, more desirable bread-making quality, and improved starch hydrolytic properties, which shows a great potential to use hulless barley as a health supplement.

The endosperm-specific transcription factor TaNAC019 regulates glutenin and starch accumulation and its elite allele improves wheat grain quality.

In wheat (Triticum aestivum L.), breeding efforts have focused intensively on improving grain yield and quality. For quality, the content and composition of seed storage proteins (SSPs) determine the elasticity of wheat dough and flour processing quality. Moreover, starch levels in seeds are associated with yield. However, little is known about the mechanisms that coordinate SSP and starch accumulation in wheat. In this study, we explored the role of the endosperm-specific NAC transcription factor TaNAC019 in coordinating SSP and starch accumulation. TaNAC019 binds to the promoters of TaGlu-1 loci, encoding high molecular weight glutenin (HMW-GS), and of starch metabolism genes. Triple knock-out mutants of all three TaNAC019 homoeologs exhibited reduced transcript levels for all SSP types and genes involved in starch metabolism, leading to lower gluten and starch contents, and in flour processing quality parameters. TaNAC019 directly activated the expression of HMW-GS genes by binding to a specific motif in their promoters and interacting with the TaGlu-1 regulator TaGAMyb. TaNAC019 also indirectly regulated the expression of TaSPA, an ortholog of maize Opaque2 that activates SSP accumulation. Therefore, TaNAC019 regulation of starch- and SSP-related genes has key roles in wheat grain quality. Finally, we identified an elite allele (TaNAC019-BI) associated with flour processing quality, providing a candidate gene for breeding wheat with improved quality.

Novel parameters characterizing size distribution of A and B starch granules in the gluten network: Effects on dough stability in bread wheat.

Our study on six wheat genotypes has revealed strong interaction between gluten and starch to affect dough stability. To establish gluten-starch interaction and its roles in dough stability, we randomly selected 16 wheat genotypes and investigated the physicochemical properties of gluten and starch. The manner in which the starch granules occupied available space in gluten network was quantitatively analyzed using gluten lacunarity and proportion of different sized A-type and B-type starch granules. Positive correlations were found between the morphological attributes (B/A/Lacunarity, B/Lacunarity) and dough stability. The correlation coefficient between B/A/Lacunarity and dough stability was highest, followed by the percentage of unextractable polymeric protein (UPP%), B/Lacunarity and dough stability. Dough mixing properties were strongly affected by gluten-starch interactions, as indicated by novel parameters. Whereas the effect of gluten on its own did not provide any evidence to suggest its concrete role in dough mixing properties because of the various genetic backgrounds.

Overexpression of GmNFYA5 confers drought tolerance to transgenic Arabidopsis and soybean plants.

BACKGROUND: Crop productivity is challenged by abiotic stresses, among which drought stress is the most common. NF-Y genes, especially NF-YA genes, regulate tolerance to abiotic stress. RESULTS: Soybean NF-Y gene GmNFYA5 was identified to have the highest transcript level among all 21 NF-YA genes in soybean (Glycine max L.) under drought stress. Drought-induced transcript of GmNFYA5 was suppressed by the ABA synthesis inhibitor naproxen (NAP). GmNFYA5 transcript was detected in various tissues at vegetative and reproductive growth stages with higher levels in roots and leaves than in other tissues, which was consist with the GmNFYA5 promoter: GUS fusion assay. Overexpression of GmNFYA5 in transgenic Arabidopsis plants caused enhanced drought tolerance in seedlings by decreasing stomatal aperture and water loss from leaves. Overexpression and suppression of GmNFYA5 in soybean resulted in increased and decreased drought tolerance, respectively, relative to plants with an empty vector (EV). Transcript levels of ABA-dependent genes (ABI2, ABI3, NCED3, LEA3, RD29A, P5CS1, GmWRKY46, GmNCED2 and GmbZIP1) and ABA-independent genes (DREB1A, DREB2A, DREB2B, GmDREB1, GmDREB2 and GmDREB3) in transgenic plants overexpressing GmNFYA5 were higher than those of wild-type plants under drought stress; suppression of GmNFYA5 transcript produced opposite results. GmNFYA5 probably regulated the transcript abundance of GmDREB2 and GmbZIP1 by binding to the promoters in vivo. CONCLUSIONS: Our results suggested that overexpression of GmNFYA5 improved drought tolerance in soybean via both ABA-dependent and ABA-independent pathways.

Genomic Analysis of Stress Associated Proteins in Soybean and the Role of GmSAP16 in Abiotic Stress Responses in Arabidopsis and Soybean.

Stress associated proteins (SAPs) containing A20/AN1 zinc finger domains have emerged as novel regulators of stress responses. In this study, 27 SAP genes were identified in soybean. The phylogenetic relationships, exon-intron structure, domain structure, chromosomal localization, putative cis-acting elements, and expression patterns of SAPs in various tissues under abiotic stresses were analyzed. Among the soybean SAP genes, GmSAP16 was significantly induced by water deficit stress, salt, and abscisic acid (ABA) and selected for further analysis. GmSAP16 was located in the nucleus and cytoplasm. The overexpression of GmSAP16 in Arabidopsis improved drought and salt tolerance at different developmental stages and increased ABA sensitivity, as indicated by delayed seed germination and stomatal closure. The GmSAP16 transgenic Arabidopsis plants had a higher proline content and a lower water loss rate and malondialdehyde (MDA) content than wild type (WT) plants in response to stresses. The overexpression of GmSAP16 in soybean hairy roots enhanced drought and salt tolerance of soybean seedlings, with higher proline and chlorophyll contents and a lower MDA content than WT. RNA inference (RNAi) of GmSAP16 increased stress sensitivity. Stress-related genes, including GmDREB1B;1, GmNCED3, GmRD22, GmDREB2, GmNHX1, and GmSOS1, showed significant expression alterations in GmSAP16-overexpressing and RNAi plants under stress treatments. These results indicate that soybean SAP genes play important roles in abiotic stress responses.

Functional Analysis of the Soybean GmCDPK3 Gene Responding to Drought and Salt Stresses.

Plants have a series of response mechanisms to adapt when they are subjected to external stress. Calcium-dependent protein kinases (CDPKs) in plants function against a variety of abiotic stresses. We screened 17 CDPKs from drought- and salt-induced soybean transcriptome sequences. The phylogenetic tree divided CDPKs of rice, Arabidopsis and soybean into five groups (I-V). Cis-acting element analysis showed that the 17 CDPKs contained some elements associated with drought and salt stresses. Quantitative real-time PCR (qRT-PCR) analysis indicated that the 17 CDPKs were responsive after different degrees of induction under drought and salt stresses. GmCDPK3 was selected as a further research target due to its high relative expression. The subcellular localization experiment showed that GmCDPK3 was located on the membrane of Arabidopsis mesophyll protoplasts. Overexpression of GmCDPK3 improved drought and salt resistance in Arabidopsis. In the soybean hairy roots experiment, the leaves of GmCDPK3 hairy roots with RNA interference (GmCDPK3-RNAi) soybean lines were more wilted than those of GmCDPK3 overexpression (GmCDPK3-OE) soybean lines after drought and salt stresses. The trypan blue staining experiment further confirmed that cell membrane damage of GmCDPK3-RNAi soybean leaves was more severe than in GmCDPK3-OE soybean lines. In addition, proline (Pro) and chlorophyll contents were increased and malondialdehyde (MDA) content was decreased in GmCDPK3-OE soybean lines. On the contrary, GmCDPK3-RNAi soybean lines had decreased Pro and chlorophyll content and increased MDA. The results indicate that GmCDPK3 is essential in resisting drought and salt stresses.

Physicochemical properties of starch in relation to rheological properties of wheat dough (Triticum aestivum L.).

Wheat dough has been considered as a complex blend where gluten forms the continuous reticular skeleton and starch granules act as filling particles. The effect of starch on dough behaviors is not clear and the mechanism of starch affecting dough properties needs to be revealed. In this study, the micro-structure and physiochemical properties of starch from six wheat varieties (lines) with different dough properties were investigated, and the rheological properties of wheat dough were determined. Six varieties with significant different starch properties perform various dough behaviors, among which Xinmai 26 with preeminent dough quality has the highest amylose content, B-type starch granule content, short-range ordered degree and starch swelling power but lowest relative crystallinity and gelatinization enthalpy of starch. The findings indicate that starch physicochemical properties also influence the dough behaviors and provide helpful information for demonstrating the effects of starch on dough properties in the protein-starch matrix.

Characterization, identification and evaluation of a set of wheat-Aegilops comosa chromosome lines.

This study characterized and evaluated a set of wheat-Aegilops comosa introgression lines, including six additions and one substitution. A total of 47 PLUG markers and a set of cytogenetic markers specific for Ae. comosa chromosomes were established after screening 526 PLUG primer pairs and performing FISH using oligonucleotides as probes. Marker analysis confirmed that these lines were wheat-Ae. comosa 2M-7M addition lines and a 6M(6A) substitution line. The molecular and cytogenetic markers developed herein could be used to trace Ae. comosa chromatin in wheat background. In order to evaluate the breeding value of the material, disease resistance tests and agronomical trait investigations were carried out on these alien chromosome introgression lines. Disease resistance tests showed that chromosomes 2M and 7M of Ae. comosa might harbor new stripe rust and powdery mildew resistance genes, respectively, therefore, they could be used as resistance sources for wheat breeding. Investigations into agronomical traits showed that all chromosomes 2M to 7M had detrimental effects on the agronomic performance of wheat, therefore, the selection of plants with relatively negative effects should be avoided when inducing wheat-A. comosa chromosome translocations using chromosome engineering procedures.

Genome-wide analysis and identification of the low potassium stress responsive gene SiMYB3 in foxtail millet (Setariaitalica L.).

BACKGROUND: Potassium (K) is essential to plant growth and development. Foxtail millet (Setaria italic L.) is an important fodder grain crop in arid and semi-arid regions of Asia and Africa because of its strong tolerance to drought and barren stresses. The molecular mechanisms of physiological and biochemical responses and regulations to various abiotic stresses such as low potassium conditions in foxtail millet are not fully understood, which hinders the research and exploitation of this valuable resource. RESULTS: In this research, we demonstrated that the millet variety Longgu 25 was the most insensitive variety to low potassium stress among other five varieties. The transcriptome analysis of Longgu 25 variety revealed a total of 26,192 and 26,849 genes from the K+-deficient and normal transcriptomic libraries by RNA-seq, respectively. A total of 1982 differentially expressed genes (DEGs) were identified including 866 up-regulated genes and 1116 down-regulated genes. We conducted a comparative analysis of these DEGs under low-K+ stress conditions and discovered 248 common DEGs for potassium deprivation among foxtail millet, rice and Arabidopsis. Further Gene Ontology (GO) enrichment analysis identified a series of candidate genes that may involve in K+-deficient response and in intersection of molecular functions among foxtail millet, rice and Arabidopsis. The expression profiles of randomly selected 18 candidate genes were confirmed as true DEGs with RT-qPCR. Furthermore, one of the 18 DEGs, SiMYB3, is specifically expressed only in the millet under low-K+ stress conditions. Overexpression of SiMYB3 promoted the main root elongation and improved K+ deficiency tolerance in transgenic Arabidopsis plants. The fresh weight of the transgenic plants was higher, the primary root length was longer and the root surface-area was larger than those of control plants after K+ deficiency treatments. CONCLUSIONS: This study provides a global view of transcriptomic resources relevant to the K+-deficient tolerance in foxtail millet, and shows that SiMYB3 is a valuable genetic resource for the improvement of K+ deficiency tolerance in foxtail millet.

Identification and Expression Analysis of Wheat TaGF14 Genes.

The 14-3-3 gene family members play key roles in various cellular processes. However, little is known about the numbers and roles of 14-3-3 genes in wheat. The aims of this study were to identify TaGF14 numbers in wheat by searching its whole genome through blast, to study the phylogenetic relationships with other plant species and to discuss the functions of TaGF14s. The results showed that common wheat harbored 20 TaGF14 genes, located on wheat chromosome groups 2, 3, 4, and 7. Out of them, eighteen TaGF14s are non-ε proteins, and two wheat TaGF14 genes, TaGF14i and TaGF14f, are ε proteins. Phylogenetic analysis indicated that these genes were divided into six clusters: cluster 1 (TaGF14d, TaGF14g, TaGF14j, TaGF14h, TaGF14c, and TaGF14n); cluster 2 (TaGF14k); cluster 3 (TaGF14b, TaGF14l, TaGF14m, and TaGF14s); cluster 4 (TaGF14a, TaGF14e, and TaGF14r); cluster 5 (TaGF14i and TaGF14f); and cluster 6 (TaGF14o, TaGF14p, TaGF14q, and TaGF14t). Tissue-specific gene expressions suggested that all TaGF14s were likely constitutively expressed, except two genes, i.e., TaGF14p and TaGF14f. And the highest amount of TaGF14 transcripts were observed in developing grains at 20 days post anthesis (DPA), especially for TaGF14j and TaGF14l. After drought stress, five genes, i.e., TaGF14c, TaGF14d, TaGF14g, TaGF14h, and TaGF14j, were up-regulated expression under drought stress for both 1 and 6 h, suggesting these genes played vital role in combating against drought stress. However, all the TaGF14s were down-regulated expression under heat stress for both 1 and 6 h, indicating TaGF14s may be negatively associated with heat stress by reducing the expression to combat heat stress or through other pathways. These results suggested that cluster 1, e.g., TaGF14j, may participate in the whole wheat developing stages, e.g., grain-filling (starch biosynthesis) and may also participate in combating against drought stress. Subsequently, a homolog of TaGF14j, TaGF14-JM22, were cloned by RACE and used to validate its function. Immunoblotting results showed that TaGF14-JM22 protein, closely related to TaGF14d, TaGF14g, and TaGF14j, can interact with AGP-L, SSI, SSII, SBEIIa, and SBEIIb in developing grains, suggesting that TaGF14s located on group 4 may be involved in starch biosynthesis. Therefore, it is possible to develop starch-rich wheat cultivars by modifying TaGF14s.