A Targeted and an Untargeted Metabolomics Approach to Study the Phytochemicals of Tomato Cultivars Grown Under Different Salinity Conditions.

In this study, we evaluated the effect of increasing the salinity of irrigation water on the metabolic content and profiles of two tomato cultivars ('Jaune Flamme' (JF) and 'Red Pear' (RP)) using targeted and untargeted metabolomics approaches. Irrigation of tomato plants was performed with four different salt concentrations provided by chloride (treatment 1) and sulfate (treatment 2) salts. Targeted analysis of the methanolic extract resulted in the identification of nine major polyphenols. Among them, chlorogenic acid, rutin, and naringenin were the prominent compounds in both cultivars. In addition, the quantification of 18 free amino acids from both tomato cultivars showed that different salinity treatments significantly enhanced the levels of glutamine, glutamic acid, and γ-aminobutyric acid (GABA). Using the untargeted metabolomic approach, we identified 129 putative metabolites encompassing a diverse array of phytochemicals including polyphenols, organic acids, lipids, sugars, and amino acids. Principal component analysis (PCA) of mass spectral data acquired under positive and negative ionization modes showed a clear separation between the two cultivars. However, only positive ionization showed separation among different salinity treatments. Unsupervised and supervised learning algorithms were applied to mine the generated data and to pinpoint metabolites different from the two cultivars. These findings suggest that different salinity conditions significantly influenced the accumulation of phytochemicals in tomato cultivars. This study will help tomato breeding programs to develop value-added tomato cultivars under varying environmental conditions.

miR828a-CsMYB114 Module Negatively Regulates the Biosynthesis of Theobromine in Camellia sinensis.

Theobromine is an important quality component in tea plants (Camellia sinensis), which is produced from 7-methylxanthine by theobromine synthase (CsTbS), the key rate-limiting enzyme in theobromine biosynthetic pathway. Our transcriptomics and widely targeted metabolomics analyses suggested that CsMYB114 acted as a potential hub gene involved in the regulation of theobromine biosynthesis. The inhibition of CsMYB114 expression using antisense oligonucleotides (ASO) led to a 70.21% reduction of theobromine level in leaves of the tea plant, which verified the involvement of CsMYB114 in theobromine biosynthesis. Furthermore, we found that CsMYB114 was located in the nucleus of the cells and showed the characteristic of a transcription factor. The dual luciferase analysis, a yeast one-hybrid assay, and an electrophoretic mobility shift assay (EMSA) showed that CsMYB114 activated the transcription of CsTbS, through binding to CsTbS promoter. In addition, a microRNA, miR828a, was identified that directly cleaved the mRNA of CsMYB114. Therefore, we conclude that CsMYB114, as a transcription factor of CsTbS, promotes the production of theobromine, which is inhibited by miR828a through cleaving the mRNA of CsMYB114.

Understanding the salt overly sensitive pathway in Prunus: Identification and characterization of NHX, CIPK, and CBL genes.

Salinity is a major abiotic stress factor that can significantly impact crop growth, and productivity. In response to salt stress, the plant Salt Overly Sensitive (SOS) signaling pathway regulates the homeostasis of intracellular sodium ion concentration. The SOS1, SOS2, and SOS3 genes play critical roles in the SOS pathway, which belongs to the members of Na+/H+ exchanger (NHX), CBL-interacting protein kinase (CIPK), and calcineurin B-like (CBL) gene families, respectively. In this study, we performed genome-wide identifications and phylogenetic analyses of NHX, CIPK, and CBL genes in six Rosaceae species: Prunus persica, Prunus dulcis, Prunus mume, Prunus armeniaca, Pyrus ussuriensis × Pyrus communis, and Rosa chinensis. NHX, CIPK, and CBL genes of Arabidopsis thaliana were used as controls for phylogenetic analyses. Our analysis revealed the lineage-specific and adaptive evolutions of Rosaceae genes. Our observations indicated the existence of two primary classes of CIPK genes: those that are intron-rich and those that are intron-less. Intron-rich CIPKs in Rosaceae and Arabidopsis can be traced back to algae CIPKs and CIPKs found in early plants, suggesting that intron-less CIPKs evolved from their intron-rich counterparts. This study identified one gene for each member of the SOS signaling pathway in P. persica: PpSOS1, PpSOS2, and PpSOS3. Gene expression analyses indicated that all three genes of P. persica were expressed in roots and leaves. Yeast two-hybrid-based protein-protein interaction analyses revealed a direct interaction between PpSOS3 and PpSOS2; and between PpSOS2 and PpSOS1C-terminus region. Our findings indicate that the SOS signaling pathway is highly conserved in P. persica.

Salinity stress tolerance prediction for biomass-related traits in maize (Zea mays L.) using genome-wide markers.

Maize (Zea mays L.) is the third most important cereal crop after rice (Oryza sativa) and wheat (Triticum aestivum). Salinity stress significantly affects vegetative biomass and grain yield and, therefore, reduces the food and silage productivity of maize. Selecting salt-tolerant genotypes is a cumbersome and time-consuming process that requires meticulous phenotyping. To predict salt tolerance in maize, we estimated breeding values for four biomass-related traits, including shoot length, shoot weight, root length, and root weight under salt-stressed and controlled conditions. A five-fold cross-validation method was used to select the best model among genomic best linear unbiased prediction (GBLUP), ridge-regression BLUP (rrBLUP), extended GBLUP, Bayesian Lasso, Bayesian ridge regression, BayesA, BayesB, and BayesC. Examination of the effect of different marker densities on prediction accuracy revealed that a set of low-density single nucleotide polymorphisms obtained through filtering based on a combination of analysis of variance and linkage disequilibrium provided the best prediction accuracy for all the traits. The average prediction accuracy in cross-validations ranged from 0.46 to 0.77 across the four derived traits. The GBLUP, rrBLUP, and all Bayesian models except BayesB demonstrated comparable levels of prediction accuracy that were superior to the other modeling approaches. These findings provide a roadmap for the deployment and optimization of genomic selection in breeding for salt tolerance in maize.

Plant phase extraction: A method for enhanced discovery of the RNA-binding proteome and its dynamics in plants.

RNA-binding proteins (RBPs) play critical roles in posttranscriptional gene regulation. Current methods of systematically profiling RBPs in plants have been predominantly limited to proteins interacting with polyadenylated (poly(A)) RNAs. We developed a method called plant phase extraction (PPE), which yielded a highly comprehensive RNA-binding proteome (RBPome), uncovering 2,517 RBPs from Arabidopsis (Arabidopsis thaliana) leaf and root samples with a highly diverse array of RNA-binding domains. We identified traditional RBPs that participate in various aspects of RNA metabolism and a plethora of nonclassical proteins moonlighting as RBPs. We uncovered constitutive and tissue-specific RBPs essential for normal development and, more importantly, revealed RBPs crucial for salinity stress responses from a RBP-RNA dynamics perspective. Remarkably, 40% of the RBPs are non-poly(A) RBPs that were not previously annotated as RBPs, signifying the advantage of PPE in unbiasedly retrieving RBPs. We propose that intrinsically disordered regions contribute to their nonclassical binding and provide evidence that enzymatic domains from metabolic enzymes have additional roles in RNA binding. Taken together, our findings demonstrate that PPE is an impactful approach for identifying RBPs from complex plant tissues and pave the way for investigating RBP functions under different physiological and stress conditions at the posttranscriptional level.

Comparative transcriptome analysis reveals key pathways and genes involved in trichome development in tea plant (Camellia sinensis).

Trichomes, which develop from epidermal cells, are considered one of the important characteristics of the tea plant [Camellia sinensis (L.) O. Kuntze]. Many nutritional and metabolomic studies have indicated the important contributions of trichomes to tea products quality. However, understanding the regulation of trichome formation at the molecular level remains elusive in tea plants. Herein, we present a genome-wide comparative transcriptome analysis between the hairless Chuyeqi (CYQ) with fewer trichomes and the hairy Budiaomao (BDM) with more trichomes tea plant genotypes, toward the identification of biological processes and functional gene activities that occur during trichome development. In the present study, trichomes in both cultivars CYQ and BDM were unicellular, unbranched, straight, and soft-structured. The density of trichomes was the highest in the bud and tender leaf periods. Further, using the high-throughput sequencing method, we identified 48,856 unigenes, of which 31,574 were differentially expressed. In an analysis of 208 differentially expressed genes (DEGs) encoding transcription factors (TFs), five may involve in trichome development. In addition, on the basis of the Gene Ontology (GO) annotation and the weighted gene co-expression network analysis (WGCNA) results, we screened several DEGs that may contribute to trichome growth, including 66 DEGs related to plant resistance genes (PRGs), 172 DEGs related to cell wall biosynthesis pathway, 29 DEGs related to cell cycle pathway, and 45 DEGs related to cytoskeleton biosynthesis. Collectively, this study provided high-quality RNA-seq information to improve our understanding of the molecular regulatory mechanism of trichome development and lay a foundation for additional trichome studies in tea plants.

Comparative Transcriptome Analysis of Agrobacterium tumefaciens Reveals the Molecular Basis for the Recalcitrant Genetic Transformation of Camellia sinensis L.

Tea (Camellia sinensis L.), an important economic crop, is recalcitrant to Agrobacterium-mediated transformation (AMT), which has seriously hindered the progress of molecular research on this species. The mechanisms leading to low efficiency of AMT in tea plants, related to the morphology, growth, and gene expression of Agrobacterium tumefaciens during tea-leaf explant infection, were compared to AMT of Nicotiana benthamiana leaves in the present work. Scanning electron microscopy (SEM) images showed that tea leaves induced significant morphological aberrations on bacterial cells and affected pathogen-plant attachment, the initial step of a successful AMT. RNA sequencing and transcriptomic analysis on Agrobacterium at 0, 3 and 4 days after leaf post-inoculation resulted in 762, 1923 and 1656 differentially expressed genes (DEGs) between the tea group and the tobacco group, respectively. The expressions of genes involved in bacterial fundamental metabolic processes, ATP-binding cassette (ABC) transporters, two-component systems (TCSs), secretion systems, and quorum sensing (QS) systems were severely affected in response to the tea-leaf phylloplane. Collectively, these results suggest that compounds in tea leaves, especially gamma-aminobutyrate (GABA) and catechins, interfered with plant-pathogen attachment, essential minerals (iron and potassium) acquisition, and quorum quenching (QQ) induction, which may have been major contributing factors to hinder AMT efficiency of the tea plant.

Deciphering Molecular Mechanisms Involved in Salinity Tolerance in Guar (Cyamopsis tetragonoloba (L.) Taub.) Using Transcriptome Analyses.

Guar is a commercially important legume crop known for guar gum. Guar is tolerant to various abiotic stresses, but the mechanisms involved in its salinity tolerance are not well established. This study aimed to understand molecular mechanisms of salinity tolerance in guar. RNA sequencing (RNA-Seq) was employed to study the leaf and root transcriptomes of salt-tolerant (Matador) and salt-sensitive (PI 340261) guar genotypes under control and salinity. Our analyses identified a total of 296,114 unigenes assembled from 527 million clean reads. Transcriptome analysis revealed that the gene expression differences were more pronounced between salinity treatments than between genotypes. Differentially expressed genes associated with stress-signaling pathways, transporters, chromatin remodeling, microRNA biogenesis, and translational machinery play critical roles in guar salinity tolerance. Genes associated with several transporter families that were differentially expressed during salinity included ABC, MFS, GPH, and P-ATPase. Furthermore, genes encoding transcription factors/regulators belonging to several families, including SNF2, C2H2, bHLH, C3H, and MYB were differentially expressed in response to salinity. This study revealed the importance of various biological pathways during salinity stress and identified several candidate genes that may be used to develop salt-tolerant guar genotypes that might be suitable for cultivation in marginal soils with moderate to high salinity or using degraded water.

Morphological, physiological, biochemical, and transcriptome studies reveal the importance of transporters and stress signaling pathways during salinity stress in Prunus.

The almond crop has high economic importance on a global scale, but its sensitivity to salinity stress can cause severe yield losses. Salt-tolerant rootstocks are vital for crop economic feasibility under saline conditions. Two commercial rootstocks submitted to salinity, and evaluated through different parameters, had contrasting results with the survival rates of 90.6% for 'Rootpac 40' (tolerant) and 38.9% for 'Nemaguard' (sensitive) under salinity (Electrical conductivity of water = 3 dS m-1). Under salinity, 'Rootpac 40' accumulated less Na and Cl and more K in leaves than 'Nemaguard'. Increased proline accumulation in 'Nemaguard' indicated that it was highly stressed by salinity compared to 'Rootpac 40'. RNA-Seq analysis revealed that a higher degree of differential gene expression was controlled by genotype rather than by treatment. Differentially expressed genes (DEGs) provided insight into the regulation of salinity tolerance in Prunus. DEGs associated with stress signaling pathways and transporters may play essential roles in the salinity tolerance of Prunus. Some additional vital players involved in salinity stress in Prunus include CBL10, AKT1, KUP8, Prupe.3G053200 (chloride channel), and Prupe.7G202700 (mechanosensitive ion channel). Genetic components of salinity stress identified in this study may be explored to develop new rootstocks suitable for salinity-affected regions.

Linking genetic determinants with salinity tolerance and ion relationships in eggplant, tomato and pepper.

The Solanaceae family includes commercially important vegetable crops characterized by their relative sensitivity to salinity. Evaluation of 8 eggplant (Solanum melongena), 7 tomato (Solanum lycopersicum), and 8 pepper (Capsicum spp.) heirloom cultivars from different geographic regions revealed significant variation in salt tolerance. Relative fruit yield under salt treatment varied from 52 to 114% for eggplant, 56 to 84% for tomato, and 52 to 99% for pepper. Cultivars from all three crops, except Habanero peppers, restricted Na transport from roots to shoots under salinity. The high salt tolerance level showed a strong association with low leaf Na concentration. Additionally, the leaf K-salinity/K-control ratio was critical in determining the salinity tolerance of a genotype. Differences in relative yield under salinity were regulated by several component traits, which was consistent with the gene expression of relevant genes. Gene expression analyses using 12 genes associated with salt tolerance showed that, for eggplant and pepper, Na+ exclusion was a vital component trait, while sequestration of Na+ into vacuoles was critical for tomato plants. The high variability for salt tolerance found in heirloom cultivars helped characterize genotypes based on component traits of salt tolerance and will enable breeders to increase the salt tolerance of Solanaceae cultivars.

Transcriptional profiling of two contrasting genotypes uncovers molecular mechanisms underlying salt tolerance in alfalfa.

Alfalfa is an important forage crop that is moderately tolerant to salinity; however, little is known about its salt-tolerance mechanisms. We studied root and leaf transcriptomes of a salt-tolerant (G03) and a salt-sensitive (G09) genotype, irrigated with waters of low and high salinities. RNA sequencing led to 1.73 billion high-quality reads that were assembled into 418,480 unigenes; 35% of which were assigned to 57 Gene Ontology annotations. The unigenes were assigned to pathway databases for understanding high-level functions. The comparison of two genotypes suggested that the low salt tolerance index for transpiration rate and stomatal conductance of G03 compared to G09 may be due to its reduced salt uptake under salinity. The differences in shoot biomass between the salt-tolerant and salt-sensitive lines were explained by their differential expressions of genes regulating shoot number. Differentially expressed genes involved in hormone-, calcium-, and redox-signaling, showed treatment- and genotype-specific differences and led to the identification of various candidate genes involved in salinity stress, which can be investigated further to improve salinity tolerance in alfalfa. Validation of RNA-seq results using qRT-PCR displayed a high level of consistency between the two experiments. This study provides valuable insight into the molecular mechanisms regulating salt tolerance in alfalfa.

Isolation and characterization of Salt Overly Sensitive family genes in spinach.

The Salt Overly Sensitive (SOS) pathway regulates intracellular sodium ion homeostasis as a salt-stress response in plants. This pathway involves three main genes designated as SOS1, SOS2 and SOS3, which are members of the Na+ /H+ exchanger (NHX), CBL-interacting protein kinase (CIPK) and Calcineurin B-like (CBL) gene families, respectively. To identify and characterize SOS genes in spinach (Spinacia oleracea), a species of the Amaranthaceae family, we conducted genome-wide identification and phylogenetic analyses of NHX, CIPK and CBL genes from four Amaranthaceae species, Arabidopsis and rice. Most Amaranthaceae genes exhibited orthologous relationships with Arabidopsis and/or rice, except a clade of Vac-type Amaranthaceae NHX genes. Phylogenetic analyses also revealed gene gain/loss events in Amaranthaceae species and the intron-less to intron-rich evolution of CIPK genes. A bacterial protein-rooted CIPK tree allowed naming most of the phylogenetic clades based on their evolutionary history. Single S. oleracea (So) SOS1, SOS2 and SOS3 proteins were identified. Direct protein-protein interaction was observed between SoSOS2 and SoSOS3 but not between SoSOS2 and SoSOS1 based on yeast two-hybrid assay. This may suggest distinct modes of action of spinach SOS proteins compared to Arabidopsis SOS proteins. Unlike SoSOS1 and SoSOS2, which were expressed at similar or higher levels in leaves than roots, SoSOS3 expression was significantly higher in roots than leaves, suggesting its greater importance in roots. The expression of SoSOS3 was upregulated in both roots and leaves under salinity compared to the control; however, SoSOS1 was only upregulated in roots. Thus, this study demonstrated the conservation of SOS pathway genes in spinach and also highlighted the complexity of SOS signaling in Amaranthaceae species.

Germination and Growth of Spinach under Potassium Deficiency and Irrigation with High-Salinity Water.

Information is scarce on the interaction of mineral deficiency and salinity. We evaluated two salt-tolerant spinach cultivars under potassium (K) doses (0.07, 0.15, 0.3, and 3.0 mmolc L-1) and saline irrigation (5, 30, 60, 120, and 160 mmolc L-1 NaCl) during germination and growth. There was no interaction between salinity and K. Salinity decreased germination percent (GP), not always significantly, and drastically reduced seedling biomass. 'Raccoon' significantly increased GP at 60 mmolc L-1 while 'Gazelle' maintained GP up to 60 or 120 mmolc L-1. After 50 days under saline irrigation, shoot biomass increased significantly at 30 and 60 mmolc L-1 at the lowest K dose but, in general, neither salinity nor K dose affected shoot biomass, suggesting that salinity supported plant growth at the most K-deficient dose. Salinity did not affect shoot N, P, or K but significantly reduced Ca, Mg, and S, although plants had no symptoms of salt toxicity or mineral deficiency. Although spinach seedlings are more sensitive to salt stress, plants adjusted to salinity with time. Potassium requirement for spinach growth was less than the current crop recommendation, allowing its cultivation with waters of moderate to high salinity without considerable reduction in yield, appearance, or mineral composition.

Linking diverse salinity responses of 14 almond rootstocks with physiological, biochemical, and genetic determinants.

Fourteen commercial almond rootstocks were tested under five types of irrigation waters to understand the genetic, physiological, and biochemical bases of salt-tolerance mechanisms. Treatments included control (T1) and four saline water treatments dominant in sodium-sulfate (T2), sodium-chloride (T3), sodium-chloride/sulfate (T4), and calcium/magnesium-chloride/sulfate (T5). T3 caused the highest reduction in survival rate and trunk diameter, followed by T4 and T2, indicating that Na and, to a lesser extent, Cl were the most toxic ions to almond rootstocks. Peach hybrid (Empyrean 1) and peach-almond hybrids (Cornerstone, Bright's Hybrid 5, and BB 106) were the most tolerant to salinity. Rootstock's performance under salinity correlated highly with its leaf Na and Cl concentrations, indicating that Na+ and Cl- exclusion is crucial for salinity tolerance in Prunus. Photosynthetic rate correlated with trunk diameter and proline leaf ratio (T3/T1) significantly correlated with the exclusion of Na+ and Cl-, which directly affected the survival rate. Expression analyses of 23 genes involved in salinity stress revealed that the expression differences among genotypes were closely associated with their performance under salinity. Our genetic, molecular, and biochemical analyses allowed us to characterize rootstocks based on component traits of the salt-tolerance mechanisms, which may facilitate the development of highly salt-tolerant rootstocks.

Spinach Plants Favor the Absorption of K+ over Na+ Regardless of Salinity, and May Benefit from Na+ When K+ is Deficient in the Soil.

Two spinach (Spinacea oleracea L.) cultivars were evaluated for their response to deficient (0.25 mmolc L-1 or 0.25 K) and sufficient (5.0 mmolc L-1 or 5.0 K) potassium (K) levels combined with salinities of 5, 30, 60, 90, and 120 mmolc L-1 NaCl. Plants substituted K for Na proportionally with salinity within each K dose. Plants favored K+ over Na+, regardless of salinity, accumulating significantly less Na at 5.0 K than at 0.25 K. Salinity had no effect on N, P, and K shoot accumulation, suggesting that spinach plants can maintain NPK homeostasis even at low soil K. Ca and Mg decreased with salinity, but plants showed no deficiency. There was no Na+ to K+ or Cl- to NO3- competition, and shoot biomass decrease was attributed to excessive NaCl accumulation. Overall, 'Raccoon' and 'Gazelle' biomasses were similar regardless of K dose but 'Raccoon' outproduced 'Gazelle' at 5.0 K at the two highest salinity levels, indicating that 'Raccoon' may outperform 'Gazelle' at higher NaCl concentrations. At low K, Na may be required by 'Raccoon', but not 'Gazelle'. This study suggested that spinach can be cultivated with recycled waters of moderate salinity, and less potassium than recommended, leading to savings on crop input and decreasing crop environmental footprint.

Characterization of natural genetic variation identifies multiple genes involved in salt tolerance in maize.

Progressive decline in irrigation water is forcing farmers to use brackish water which increases soil salinity and exposes the crop plants to salinity. Maize, one of the most important crops, is sensitive to salinity. Salt tolerance is a complex trait controlled by a number of physiological and biochemical processes. Scant information is available on the genetic architecture of salt tolerance in maize. We evaluated 399 inbred lines for six early vigor shoot and root traits upon exposure of 18-day seedlings to salinity (ECiw = 16 dS m-1) stress. Contrasting response of shoot and root growth to salinity indicated a meticulous reprogramming of resource partitioning by the plants to cope with the stress. The genomic analysis identified 57 single nucleotide polymorphisms (SNP) associated with early vigor traits. Candidate genes systematically associated with each SNP include both previously known and novel genes. Important candidates include a late embryogenesis protein, a divalent ion symporter, a proton extrusion protein, an RNA-binding protein, a casein kinase 1, and an AP2/EREBP transcription factor. The late embryogenesis protein is associated with both shoot and root length, indicating a coordinated change in resource allocation upon salt stress. Identification of a casein kinase 1 indicates an important role for Ser/Thr kinases in salt tolerance. Validation of eight candidates based on expression in a salt-tolerant and a salt-sensitive inbred line supported their role in salt tolerance. The candidate genes identified in this investigation provide a foundation for dissecting genetic and molecular regulation of salt tolerance in maize and related grasses.

Mutation in a PHD-finger protein MS4 causes male sterility in soybean.

BACKGROUND: Male sterility has tremendous scientific and economic importance in hybrid seed production. Identification and characterization of a stable male sterility gene will be highly beneficial for making hybrid seed production economically feasible. In soybean, eleven male-sterile, female-fertile mutant lines (ms1, ms2, ms3, ms4, ms5, ms6, ms7, ms8, ms9, msMOS, and msp) have been identified and mapped onto various soybean chromosomes, however the causal genes responsible for male sterility are not isolated. The objective of this study was to identify and functionally characterize the gene responsible for the male sterility in the ms4 mutant. RESULTS: The ms4 locus was fine mapped to a 216 kb region, which contains 23 protein-coding genes including Glyma.02G243200, an ortholog of Arabidopsis MALE MEIOCYTE DEATH 1 (MMD1), which is a Plant Homeodomain (PHD) protein involved in male fertility. Isolation and sequencing of Glyma.02G243200 from the ms4 mutant line showed a single base insertion in the 3rd exon causing a premature stop codon resulting in truncated protein production. Phylogenetic analysis showed presence of a homolog protein (MS4_homolog) encoded by the Glyma.14G212300 gene. Both proteins were clustered within legume-specific clade of the phylogenetic tree and were likely the result of segmental duplication during the paleoploidization events in soybean. The comparative expression analysis of Ms4 and Ms4_homologs across the soybean developmental and reproductive stages showed significantly higher expression of Ms4 in early flowering (flower bud differentiation) stage than its homolog. The functional complementation of Arabidopsis mmd1 mutant with the soybean Ms4 gene produced normal stamens, successful tetrad formation, fertile pollens and viable seeds, whereas the Ms4_homolog was not able to restore male fertility. CONCLUSIONS: Overall, this is the first report, where map based cloning approach was employed to isolate and characterize a gene responsible for the male-sterile phenotype in soybean. Characterization of male sterility genes may facilitate the establishment of a stable male sterility system, highly desired for the viability of hybrid seed production in soybean. Additionally, translational genomics and genome editing technologies can be utilized to generate new male-sterile lines in other plant species.

Expression of the high-affinity K+ transporter 1 (PpHKT1) gene from almond rootstock 'Nemaguard' improved salt tolerance of transgenic Arabidopsis.

Soil salinity affects plant growth and development, which directly impact yield. Plants deploy many mechanisms to cope with, or mitigate, salt stress. One of such mechanism is to control movement of ions from root to shoot by regulating the loading of Na+ in the transpiration stream. The high-affinity K+ transporter 1 (HKT1) is known to play a role in the removal of Na+ from the xylem and bring it back to the root. As almond is a salt-sensitive crop, the rootstock plays an important role in successful almond cultivation in salt-affected regions. We currently lack knowledge on the molecular mechanisms involved in salt tolerance of almond rootstocks. In this study, we complemented the Arabidopsis athkt1 knockout mutant with HKT1 ortholog (PpHKT1) from the almond rootstock 'Nemaguard'. Arabidopsis transgenic lines that were generated in athkt1 background with the constitutive promoter (PpHKT1OE2.2) and the native promoter (PpHKT1NP6) were subjected to different salt treatments. Both transgenic lines survived salt concentrations up to 120 mM NaCl, however, the mutant athkt1 died after 18 days under 120 mM NaCl. At 90 mM NaCl, the dry weight of athkt1 decreased significantly compared to the transgenic lines. Both transgenic lines showed significantly longer lateral roots compared to the athkt1 mutant at 80 mM NaCl treatment. The transgenic lines, PpHKT1OE2.2 and PpHKTNP6 had lower electrolyte leakage and higher relative water content compared to athkt1, suggesting that transgenic plants coped well with increased salt concentration by maintaining the integrity of the membranes. The expression analyses showed that PpHKT1 was induced in PpHKT1OE2.2 and PpHKTNP6 lines under salt treatment, which confirmed that both over-expression and native expression of PpHKT1 in the Arabidopsis mutant can complement salt tolerance function.

Seasonal and Differential Sesquiterpene Accumulation in Artemisia annua Suggest Selection Based on Both Artemisinin and Dihydroartemisinic Acid may Increase Artemisinin in planta.

Commercial Artemisia annua crops are the sole source of artemisinin (ART) worldwide. Data on seasonal accumulation and peak of sesquiterpenes, especially ART in commercial A. annua, is lacking while current breeding programs focus only on ART and plant biomass, but ignores dihydroartemisinic acid (DHAA) and artemisinic acid (AA). Despite past breeding successes, plants richer in ART are needed to decrease prices of artemisinin-combination therapy (ACT). Our results show that sesquiterpene concentrations vary greatly along the growing season and that sesquiterpene profiles differ widely among chemotypes. Field studies with elite Brazilian, Chinese, and Swiss germplasms established that ART peaked in vegetative plants from late August to early September, suggesting that ART is related to the photoperiod, not flowering. DHAA peaks with ART in Chinese and Swiss plants, but decreases, as ART increases, in Brazilian plants, while AA remained stable through the season in these genotypes. Chinese plants peaked at 0.9% ART, 1.6% DHAA; Brazilian plants at 0.9% ART, with less than 0.4% DHAA; Swiss plants at 0.8% ART and 1% DHAA. At single-date harvests, seeded Swiss plants produced 0.55-1.2% ART, with plants being higher in DHAA than ART; Brazilian plants produced 0.33-1.5% ART, with most having higher ART than DHAA. Elite germplasms produced from 0.02-0.43% AA, except Sandeman-UK (0.4-1.1% AA). Our data suggest that different chemotypes, high in ART and DHAA, have complementary pathways, while competing with AA. Crossing plants high in ART and DHAA may generate hybrids with higher ART than currently available in commercial germplasms. Selecting for high ART and DHAA (and low AA) can be a valuable approach for future selection and breeding to produce plants more efficient in transforming DHAA into ART in planta and during post-harvest. This novel approach could change the breeding focus of A. annua and other pharmaceutical species that produce more than one desired metabolite in the same pathway. Obtaining natural variants with high ART content will empower countries and farmers who select, improve, and cultivate A. annua as a commercial pharmaceutical crop. This selection approach could enable ART to be produced locally where it is most needed to fight malaria and other parasitic neglected diseases.

Inheritance and Genetic Mapping of the Reduced Height (Rht18) Gene in Wheat.

Short-statured plants revolutionized agriculture during the 1960s due to their ability to resist lodging, increased their response to fertilizers, and improved partitioning of assimilates which led to yield gains. Of more than 21 reduced-height (Rht) genes reported in wheat, only three-Rht-B1b, Rht-D1b, and Rht8-were extensively used in wheat breeding programs. The remaining reduced height mutants have not been utilized in breeding programs due to the lack of characterization. In the present study, we determined the inheritance of Rht18 and developed a genetic linkage map of the region containing Rht18. The height distribution of the F2 population was skewed towards the mutant parent, indicating that the dwarf allele (Rht18) is semi-dominant over the tall allele (rht18). Rht18 was mapped on chromosome 6A between markers barc146 and cfd190 with a genetic distance of 26.2 and 17.3 cM, respectively. In addition to plant height, agronomically important traits, like awns and tiller numbers, were also studied in the bi-parental population. Although the average tiller number was very similar in both parents, the F2 population displayed a normal distribution for tiller number with the majority of plants having phenotype similar to the parents. Transgressive segregation was observed for plant height and tiller number in F2 population. This study enabled us to select a semi-dwarf line with superior agronomic characteristics that could be utilized in a breeding program. The identification of SSRs associated with Rht18 may improve breeders' effectiveness in selecting desired semi-dwarf lines for developing new wheat cultivars.