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Sucrose non-fermenting 1-related protein kinase 2 (SnRK2) is a plant-specific serine/threonine kinase involved in response to adverse environmental stimuli. Previous studies showed that TaSnRK2.4 was involved in response to abiotic stresses and conferred enhanced tolerance to multiple stresses in Arabidopsis. Further experiments were performed to decipher the underlying mechanisms and discover new functions. The genomic sequences of TaSnRK2.4s locating on chromosome 3A, 3B and 3D were obtained. Sequencing identified one and 13 variations of TaSnRK2.4–3A and TaSnRK2.4-3B, respectively, but no variation was detected in TaSnRK2.4-3D. The markers 2.4AM1, 2.4BM1 and 2.4BM2 were developed based on three variations. Association analysis showed that both TaSnRK2.4-3A and TaSnRK2.4-3B were significantly associated with thousand-kernel weight (TKW), and that SNP3A-T and SNP3B-C were favorable alleles for higher TKW. Yeast two-hybrid and split luciferase assays showed that TaSnRK2.4 physically interacted with abiotic stress responsive protein TaLTP3, suggesting that TaSnRK2.4 enhanced abiotic stress tolerance by activating TaLTP3. Our studies suggested that TaSnRK2.4 have potential in improving TKW and response to abiotic stress.

Nitrate transporters (NRTs) are regulators of nitrate assimilation and transport. The genome sequences of TaNRT2L12-A, -B and -D were cloned from wheat (Triticum aestivum L.), and polymorphisms were analyzed by sequencing. TaNRT2L12-D in a germplasm population was highly conserved. However, 38 single nucleotide polymorphisms (SNPs) in TaNRT2L12-A coding region and 11 SNPs in TaNRT2L12-B coding region were detected. Two derived cleaved amplified polymorphic sequences (dCAPS) markers A-CSNP1 and A-CSNP2 were developed for TaNRT2L12-A based on SNP-351 and SNP-729, and three haplotypes were identified in the germplasm population. B-CSNP1 and B-CSNP2 were developed for TaNRT2L12-B based on SNP-237 and SNP-1227, and three haplotypes were detected in the germplasm population. Association analyses between the markers and agronomic traits in 30 environments and phenotypic comparisons revealed that A-CSNP2-A is a superior allele of shorter plant height (PH), length of penultimate internode (LPI) and peduncle length (PL), B-CSNP2-G is a superior allele of higher grain number per spike (GNS). Hap-6B-1 containing both superior alleles B-CSNP1-C and B-CSNP2-A is a superior haplotype of 1 000-grain weight (TGW). Expression analysis showed that TaNRT2L12-B is mainly expressed in the root base and regulated by nitrate. Therefore, TaNRT2L12 may be involved in nitrate transport and signaling to regulate TGW in wheat. The superior alleles and dCAPS markers of TaNRT2L12-A/B are beneficial to genetic improvement and germplasm enhancement with molecular markers-assisted selection.

The stress-associated protein (SAP) multigene family is conserved in both animals and plants. Its function in some animals and plants are known, but it is yet to be deciphered in wheat (Triticum aestivum L.). We identified the wheat gene TaSAP17-D, a member of the SAP gene family with an AN1/AN1 conserved domain. Subcellular localization indicated that TaSAP17-D localized to the nucleus, cytoplasm, and cell membrane. Expression pattern analyses revealed that TaSAP17-D was highly expressed in seedlings and was involved in NaCl response, polyethylene glycol (PEG), cold, and exogenous abscisic acid (ABA). Constitutive expression of TaSAP17-D in transgenic Arabidopsis resulted in enhanced tolerance to salt stress, confirmed by improved multiple physiological indices and significantly upregulated marker genes related to salt stress response. Our results suggest that TaSAP17-D is a candidate gene that can be used to protect crop plants from salt stress.

Stress associated proteins (SAPs) are the A20/AN1 zinc-finger proteins which confer to abiotic stresses in plants. In this study, TaSAP7-B, including two AN1 domains, was isolated from B genome of wheat (Triticum aestivum L.). Sequencing analysis on TaSAP7-B illustrated one InDel (insertion-deletion) and one SNP (single nucleotide polymorphism) in the promoter region while no diversity was observed in the coding region. On the basis of SNP in the promoter region (–260 bp), a dCAPS (derived cleaved amplified polymorphic sequences) marker SNP-260 was developed for TaSAP7-B. Using a natural population consisting of 262 wheat accessions, significant associations were detected between the marker SNP-260 and agronomic traits, such as plant height (PH), peduncle length (PL), length of penultimate internode (LPI), number of spike per plant (NSP), and 1 000-grain weight (TGW). Two genotypes were identified using marker SNP-260 in the natural population. Among them, the genotypes possessing C allele exhibited a higher TGW and shorter PH than the T genotypes. Hence, base C was considered as the superior allele. The dCAPS marker of TaSAP7-B can be instrumental for marker-assisted selection for high grain size and short plant height.

Fructans are major nonstructural carbohydrates in wheat (Triticum aestivum L.). Fructan 1-fructosyltransferase (1-FFT) is the key enzyme in fructan biosynthesis. In the present study, 96 sequence variants were detected in the 1-FFT-A1 gene among 26 wheat accessions including UR208, and 15 of them result in amino acid substitutions, forming four haplotypes. Two markers M39 and M2164 were developed based on the InDel21-39 and SNP-2164 polymorphisms to distinguish the three haplotypes in the 1-FFT-A1. 1-FFT-A1 was located on chromosome 4A using marker M2164 and was flanked by markers Xcwm27 and 6-SFT-A1. By association analysis using a natural wheat population consisted of 154 accessions, the results showed that the two markers were significantly associated with water-soluble carbohydrate (WSC) content in the lower internode stem and total stem at the early and middle grain filling stages, 1000-grain weight (TGW) at different grain filling stages and peduncle length (PLE). Comparison of the effects of three haplotypes on agronomic traits indicated that TGW, PLE and total number of spikelets per spike (TNSS) were significantly influenced by haplotypes. HapIII showed a significant positive effect on TGW, PLE and TNSS.

Lipid transfer protein (LTP) is a kind of small molecular protein, which is named for its ability to transfer lipid between cell membranes. It has been proved that the protein is involved in the responding to abiotic stresses. In this study, TaLTP-s, a genomic sequence of TaLTP was isolated from A genome of wheat (Triticum aestivum L). Sequencing analysis exhibited that there was no diversity in the coding region of TaLTP-s, but seven single nucleotide polymorphisms (SNPs) and 1 bp insertion/deletion (InDel) were detected in the promoter regions of different wheat accessions. Nucleotide diversity (π) in the region was 0.00033, and linkage disequilibrium (LD) extended over almost the entire TaLTP-s region in wheat. The dCAPS markers based on sequence variations in the promoter regions (SNP-207 and SNP-1696) were developed, and three haplotypes were identified based on those markers. Association analysis between the haplotypes and agronomic traits of natural population consisted of 262 accessions showed that three haplotypes of TaLTP-s were significantly associated with plant height (PH). Among the three haplotypes, HapIII is considered as the superior haplotype for increasing plant height in the drought stress environments. The G variance at the position of 207 bp could be a superior allele that significantly increased number of spikes per plant (NSP). The functional marker of TaLTP-s provide a tool for marker-assisted selection regarding to plant height and number of spikelet per plant in wheat.

Stress-associated protein (SAP) has functions in maintaining plant cell elongation, embryo development and response to abiotic stresses. TaSAP1-A1 , one of the Triticum aestivum SAP1 ( TaSAP1 ) members located on wheat chromosome 7A was isolated for polymorphism analysis. Hap III of TaSAP1-A1 was found significantly associated with thousand-grain weight (TGW) in multiple environments. In this study, Hap III also made a positive contribution to TGW in Population 2. The distribution of TaSAP1-A1 Hap III was tracked among varieties released in different years and geographical environments of China. The frequency of Hap III showed an increasing trend during the breeding process in two different populations. The Hap III was gradually selected and applied from 6.36% in landraces to 13.50% in modern varieties. These results exhibited that TaSAP1-A1 Hap III was positively selected during wheat breeding, which is beneficial for grain-yield improvement. The preferred Hap III was initially selected and applied in the higher latitude areas of China in accord with the long day season and longer grain filling stage in these areas. Moreover, the frequency of Hap III in recent modern varieties was still quite low (19.29–26.67%). It indicated a high application potential of TaSAP1-A1 Hap III for improving grain yield in wheat breeding.

Wheat is an important source of essential minerals for human body. Breeding wheat with high grain mineral concentration thus benefits human health. The objective of present study was to identify quantitative trait loci (QTLs) controlling grain mineral concentration and to evaluate the relation between nitrogen (N) and other essential minerals in winter wheat. Wheat grains were harvested from field experiment which conducted in China and analyzed for this purpose. Forty-three QTLs controlling grain mineral concentration and nitrogen-related traits were detected by using a double haploid (DH) population derived from winter wheat varieties Hanxuan 10 and Lumai 14. Chromosomes 4D and 5A might be very important in controlling mineral status in wheat grains. Significant positive correlations were found between grain nitrogen concentration (GNC) and nutrients Fe, Mn, Cu, Mg concentrations (FeGC, MnGC, CuGC, MgGC). Flag leaf N concentration at anthesis (FLNC) significantly and positively correlated with GNC, FeGC, MnGC, and CuGC. The study extended our knowledge on minerals in wheat grains and suggested which interactions between minerals should be considered in future breeding program.

Plant cysteine protease (CP) genes are induced by abiotic stresses such as drought, yet their functions remain largely unknown. We isolated the full-length cDNA encoding a Triticum aestivum CP gene, designated TaCP, from wheat by the rapid amplification of cDNA ends (RACE) method. Sequence analysis revealed that TaCP contains an open reading frame encoding a protein of 362 amino acids, which is 96% identical to barley cysteine protease HvSF42. The TaCP transcript level in wheat seedlings was upregulated during polyethylene glycol (PEG) stress, with a peak appearing around 12 h after treatment. TaCP expression level increased rapidly with NaCl treatment at 48 h. TaCP responded strongly to low temperature (4 degree C) treatment from 1 h post-treatment and reached a peak of about 40-fold at 72 h. However, it showed only a very slight response to abscisic acid (ABA). More than one copy of TaCP was present in each of the three genomes of hexaploid wheat and its diploid donors. TaCP fused with green fluorescent protein (GFP) was located in the plasma membrane of onion epidermis cells. Transgenic Arabidopsis plants overexpressing TaCP showed stronger drought tolerance and higher CP activity under water-stressed conditions than wild-type Arabidopsis plants. The results suggest that TaCP plays a role in tolerance to water deficit.

A full-length cDNA denominated PvP5CS for Delta(1)-pyrroline-5-carboxylate synthetase (P5CS), an enzyme involved in the biosynthesis of proline, was cloned from common bean using a candidate gene approach. PvP5CS contains an open reading frame encoding a 716 amino acid polypeptide. Sequence analysis showed that PvP5CS shares 95.1% homology in nucleotide sequence and 93.2% identity in amino acid sequence with the mothbean (Vigna aconitifolia) P5CS. The expression patterns of PvP5CS in common bean treated with drought, cold (4 degrees C), and salt (200 mM NaCl) stresses were examined using real-time quantitative PCR. These abiotic stresses caused significant up-regulation of the expression of PvP5CS in leaves. The PvP5CS mRNA transcript increased to 2.5 times the control level after 4d drought stress. A rapid up-regulation of PvP5CS, to about 16.3 times the control at 2h post-treatment was observed under salt stress. A significant increase in PvP5CS expression (11.7-fold) was detected after 2h of cold stress. The peaks of proline accumulation appeared at 8d for drought, 24h for cold and 9h for salt stress, somewhat later than the peaks of PvP5CS expression. These results suggest that PvP5CS was a stress-inducible gene regulating the accumulation of proline in plants subjected to stress. Finally, subcellular localization assays showed that the PvP5CS protein was present in the nucleus and at the plasmalemma.

Parameters of chlorophyll fluorescence kinetics (PCFKs) under drought stress condition are generally used to characterize instincts for dehydration tolerance in wheat (Triticum aestivum L.). Therefore, it is important to map quantitative trait loci (QTLs) for PCFKs in wheat genetic improvement for drought tolerance. A doubled haploid (DH) population with 150 lines, derived from a cross between two common wheat varieties, Hanxuan 10 and Lumai 14, was used to analyze the correlation between PCFKs and chlorophyll content (ChlC) and to map QTLs at the grain-filling stage under conditions of both rainfed (drought stress, DS) and well-watered (WW), respectively. QTLs for these traits were detected by QTLMapper version 1.0 based on the composite interval mapping method of the mixed-linear model. The results showed a very significant positive correlation between Fv, Fm, Fv/Fm and Fv/Fo. The correlation coefficients were generally higher under WW than under DS. Also, there was a significant or a highly significant positive correlation between Fv, Fm, Fv/Fm, Fv/Fo and ChlC. The correlation coefficients were higher in the DS group than the WW group. A total of 14 additive QTLs (nine QTLs detected under DS and five QTLs under WW) and 25 pairs of epistatic QTLs (15 pairs detected under DS and 10 pairs under WW) for PCFKs were mapped on chromosomes 6A, 7A, 1B, 3B, 4D and 7D. The contributions of additive QTLs for PCFKs to phenotype variation were from 8.40% to 72.72%. Four additive QTLs (two QTLs detected under DS and WW apiece) controlling ChlC were mapped on chromosomes 1A, 5A and 7A. The contributions of these QTLs for ChlC to phenotype variation were from 7.27% to 11.68%. Several QTL clusters were detected on chromosomes 1B, 7A and 7D, but no shared chromosomal regions for them were identified under different water regimes, indicating that these QTLs performed different expression patterns under rainfed and well-watered conditions. (Handling editor: Yong-Biao Xue)

†Background and Aims Multiple copies of genes encoding the catalytic subunit (c) of protein phosphatase 2A (PP2A) are commonly found in plants. For some of these genes, expression is up-regulated under water stress. The aim of this study was to investigate expression and characterization of TaPP2Ac-1 from Triticum aestivum, and to evaluate the effects of TaPP2Ac-1 on Nicotiana benthamiana in response to water stress. †Methods TaPP2Ac-1 cDNA was isolated from wheat by in silico identification and RT-PCR amplification. Transcript levels of TaPP2Ac-1 were examined in wheat responding to water deficit. Copy numbers of TaPP2Ac-1 in wheat genomes and subcellular localization in onion epidermal cells were studied. Enzyme properties of the recombinant TaPP2Ac-1 protein were determined. In addition, studies were carried out in tobacco plants with pCAPE2-TaPP2Ac-1 under water-deficit conditions. †Key Results TaPP2Ac-1 cDNA was cloned from wheat. Transcript levels of TaPP2Ac-1 in wheat seedlings were up-regulated under drought condition. One copy for this TaPP2Ac-1 was present in each of the three wheat genomes. TaPP2Ac-1 fused with GFP was located in the nucleus and cytoplasm of onion epidermis cells. The recombinant TaPP2Ac-1 gene was over-expressed in Escherichia coli and encoded a functional serine/threonine phosphatase. Transgenic tobacco plants over-expressing TaPP2Ac-1 exhibited stronger drought tolerance than nontransgenic tobacco plants. †Conclusions Tobacco plants with pCAPE2-TaPP2Ac-1 appeared to be resistant to water deficit, as shown by their higher capacity to maintain leaf relative water content, leaf cell-membrane stability index, water-retention ability and water use efficiency under water stress. The results suggest that the physiological role of TaPP2Ac-1 is related to drought stress response, possibly through its involvement in drought-responding signal transduction pathways.