OsPIN5b modulates rice (Oryza sativa) plant architecture and yield by changing auxin homeostasis,transport and distribution

Plant architecture attributes such as tillering, plant height and panicle size are important agronomic traits that determine rice (Oryza sativa) productivity. Here, we report that altered auxin content, transport and distribution affect these traits, and hence rice yield. Overexpression of the auxin efflux carrier‐like gene OsPIN5b causes pleiotropic effects, mainly reducing plant height, leaf and tiller number, shoot and root biomass, seed‐setting rate, panicle length and yield parameters. Conversely, reduced expression of OsPIN5b results in higher tiller number, more vigorous root system, longer panicles and increased yield. We show that OsPIN5b is an endoplasmic reticulum (ER) ‐localized protein that participates in auxin homeostasis, transport and distribution in vivo. This work describes an example of an auxin‐related gene where modulating its expression can simultaneously improve plant architecture and yield potential in rice, and reveals an important effect of hormonal signaling on these traits.     

Oligonucleotide‐directed mutagenesis for precision gene editing

Differences in gene sequences, many of which are single nucleotide polymorphisms, underlie some of the most important traits in plants. With humanity facing significant challenges to increase global agricultural productivity, there is an urgent need to accelerate the development of these traits in plants. oligonucleotide‐directed mutagenesis (ODM), one of the many tools of Cibus’ Rapid Trait Development System ( RTDS ^?) technology, offers a rapid, precise and non‐transgenic breeding alternative for trait improvement in agriculture to address this urgent need. This review explores the application of ODM as a precision genome editing technology, with emphasis on using oligonucleotides to make targeted edits in plasmid, episomal and chromosomal DNA of bacterial, fungal, mammalian and plant systems. The process of employing ODM by way of RTDS technology has been improved in many ways by utilizing a fluorescence conversion system wherein a blue fluorescent protein (BFP) can be changed to a green fluorescent protein (GFP) by editing a single nucleotide of the BFP gene (CAC→TAC; H66 to Y66). For example, dependent on oligonucleotide length, applying oligonucleotide‐mediated technology to target the BFP transgene in Arabidopsis thaliana protoplasts resulted in up to 0.05% precisely edited GFP loci. Here, the development of traits in commercially relevant plant varieties to improve crop performance by genome editing technologies such as ODM, and by extension RTDS , is reviewed.     

Approaches for enhancement of N2 fixation efficiency of chickpea (Cicer arietinum L.) under limiting nitrogen conditions

Chickpea (Cicer arietinum) is an important pulse crop in many countries in the world. The symbioses between chickpea and Mesorhizobia, which fix N₂ inside the root nodules, are of particular importance for chickpea's productivity. With the aim of enhancing symbiotic efficiency in chickpea, we compared the symbiotic efficiency of C‐15, Ch‐191 and CP‐36 strains of Mesorhizobium ciceri in association with the local elite chickpea cultivar ‘Bivanij’ as well as studied the mechanism underlying the improvement of N₂ fixation efficiency. Our data revealed that C‐15 strain manifested the most efficient N₂ fixation in comparison with Ch‐191 or CP‐36. This finding was supported by higher plant productivity and expression levels of the nifHDK genes in C‐15 nodules. Nodule specific activity was significantly higher in C‐15 combination, partially as a result of higher electron allocation to N₂ versus H⁺. Interestingly, a striking difference in nodule carbon and nitrogen composition was observed. Sucrose cleavage enzymes displayed comparatively lower activity in nodules established by either Ch‐191 or CP‐36. Organic acid formation, particularly that of malate, was remarkably higher in nodules induced by C‐15 strain. As a result, the best symbiotic efficiency observed with C‐15‐induced nodules was reflected in a higher concentration of the total and several major amino metabolites, namely asparagine, glutamine, glutamate and aspartate. Collectively, our findings demonstrated that the improved efficiency in chickpea symbiotic system, established with C‐15, was associated with the enhanced capacity of organic acid formation and the activities of the key enzymes connected to the nodule carbon and nitrogen metabolism.     

PAY1 improves plant architecture and enhances grain yield in rice

Plant architecture, a complex of the important agronomic traits that determine grain yield, is a primary target of artificial selection of rice domestication and improvement. Some important genes affecting plant architecture and grain yield have been isolated and characterized in recent decades; however, their underlying mechanism remains to be elucidated. Here, we report genetic identification and functional analysis of the PLANT ARCHITECTURE AND YIELD 1 (PAY1) gene in rice, which affects plant architecture and grain yield in rice. Transgenic plants over‐expressing PAY1 had twice the number of grains per panicle and consequently produced nearly 38% more grain yield per plant than control plants. Mechanistically, PAY1 could improve plant architecture via affecting polar auxin transport activity and altering endogenous indole‐3‐acetic acid distribution. Furthermore, introgression of PAY1 into elite rice cultivars, using marker‐assisted background selection, dramatically increased grain yield compared with the recipient parents. Overall, these results demonstrated that PAY1 could be a new beneficial genetic resource for shaping ideal plant architecture and breeding high‐yielding rice varieties.     

Genetic dissection of pre‐anthesis sub‐phase durations during the reproductive spike development of wheat

Flowering time is an important factor affecting grain yield in wheat. In this study, we divided reproductive spike development into eight sub‐phases. These sub‐phases have the potential to be delicately manipulated to increase grain yield. We measured 36 traits with regard to sub‐phase durations, determined three grain yield‐related traits in eight field environments and mapped 15 696 single nucleotide polymorphism (SNP, based on 90k Infinium chip and 35k Affymetrix chip) markers in 210 wheat genotypes. Phenotypic and genetic associations between grain yield traits and sub‐phase durations showed significant consistency (Mantel test; r = 0.5377, P < 0.001). The shared quantitative trait loci (QTLs) revealed by the genome‐wide association study suggested a close association between grain yield and sub‐phase duration, which may be attributed to effects on spikelet initiation/spikelet number (double ridge to terminal spikelet stage, DR‐TS) and assimilate accumulation (green anther to anthesis stage, GA‐AN). Moreover, we observed that the photoperiod‐sensitivity allele at the Ppd‐D1 locus on chromosome 2D markedly extended all sub‐phase durations, which may contribute to its positive effects on grain yield traits. The dwarfing allele at the Rht‐D1 (chromosome 4D) locus altered the sub‐phase duration and displayed positive effects on grain yield traits. Data for 30 selected genotypes (from among the original 210 genotypes) in the field displayed a close association with that from the greenhouse. Most importantly, this study demonstrated specific connections to grain yield in narrower time windows (i.e. the eight sub‐phases), rather than the entire stem elongation phase as a whole.     

Sex‐biased oviposition by a nursery pollinator on a gynodioecious host plant: Implications for breeding system evolution and evolution of mutualism

Dioecy, a breeding system where individual plants are exclusively male or female, has evolved repeatedly. Extensive theory describes when dioecy should arise from hermaphroditism, frequently through gynodioecy, where females and hermaphrodites coexist, and when gynodioecy should be stable. Both pollinators and herbivores often prefer the pollen‐bearing sex, with sex‐specific fitness effects that can affect breeding system evolution. Nursery pollination, where adult insects pollinate flowers but their larvae feed on plant reproductive tissues, is a model for understanding mutualism evolution but could also yield insights into plant breeding system evolution. We studied a recently established nursery pollination interaction between native Hadena ectypa moths and introduced gynodioecious Silene vulgaris plants in North America to assess whether oviposition was biased toward females or hermaphrodites, which traits were associated with oviposition, and the effect of oviposition on host plant fitness. Oviposition was hermaphrodite‐biased and associated with deeper flowers and more stems. Sexual dimorphism in flower depth, a trait also associated with oviposition on the native host plant (Silene stellata), explained the hermaphrodite bias. Egg‐receiving plants experienced more fruit predation than plants that received no eggs, but relatively few fruits were lost, and egg receipt did not significantly alter total fruit production at the plant level. Oviposition did not enhance pollination; egg‐receiving flowers usually failed to expand and produce seeds. Together, our results suggest that H. ectypa oviposition does not exert a large fitness cost on host plants, sex‐biased interactions can emerge from preferences developed on a hermaphroditic host species, and new nursery pollination interactions can arise as negative or neutral rather than as mutualistic for the plant.     

Constitutive or seed‐specific overexpression of Arabidopsis G‐protein γ subunit 3 (AGG3) results in increased seed and oil production and improved stress tolerance in Camelina sativa

Heterotrimeric G‐proteins consisting of Gα, Gβ and Gγ subunits play an integral role in mediating multiple signalling pathways in plants. A novel, recently identified plant‐specific Gγ protein, AGG3, has been proposed to be an important regulator of organ size and mediator of stress responses in Arabidopsis, whereas its potential homologs in rice are major quantitative trait loci for seed size and panicle branching. To evaluate the role of AGG3 towards seed and oil yield improvement, the gene was overexpressed in Camelina sativa, an oilseed crop of the Brassicaceae family. Analysis of multiple homozygous T4 transgenic Camelina lines showed that constitutive overexpression of AGG3 resulted in faster vegetative as well as reproductive growth accompanied by an increase in photosynthetic efficiency. Moreover, when expressed constitutively or specifically in seed tissue, AGG3 was found to increase seed size, seed mass and seed number per plant by 15%–40%, effectively resulting in significantly higher oil yield per plant. AGG3 overexpressing Camelina plants also exhibited improved stress tolerance. These observations draw a strong link between the roles of AGG3 in regulating two critical yield parameters, seed traits and plant stress responses, and reveal an effective biotechnological tool to dramatically increase yield in agricultural crops.     

A multiparental cross population for mapping QTL for agronomic traits in durum wheat (Triticum turgidum ssp. durum)

Multiparental cross designs for mapping quantitative trait loci (QTL) provide an efficient alternative to biparental populations because of their broader genetic basis and potentially higher mapping resolution. We describe the development and deployment of a recombinant inbred line (RIL) population in durum wheat (Triticum turgidum ssp. durum) obtained by crossing four elite cultivars. A linkage map spanning 2664 cM and including 7594 single nucleotide polymorphisms (SNPs) was produced by genotyping 338 RILs. QTL analysis was carried out by both interval mapping on founder haplotype probabilities and SNP bi‐allelic tests for heading date and maturity date, plant height and grain yield from four field experiments. Sixteen QTL were identified across environments and detection methods, including two yield QTL on chromosomes 2BL and 7AS, with the former mapped independently from the photoperiod response gene Ppd‐B1, while the latter overlapped with the vernalization locus VRN‐A3. Additionally, 21 QTL with environment‐specific effects were found. Our results indicated a prevalence of environment‐specific QTL with relatively small effect on the control of grain yield. For all traits, functionally different QTL alleles in terms of direction and size of genetic effect were distributed among parents. We showed that QTL results based on founder haplotypes closely matched functional alleles at known heading date loci. Despite the four founders, only 2.1 different functional haplotypes were estimated per QTL, on average. This durum wheat population provides a mapping resource for detailed genetic dissection of agronomic traits in an elite background typical of breeding programmes.     

Monitoring codling moth (Lepidoptera: Tortricidae) in orchards treated with pear ester and sex pheromone combo dispensers

Monitoring codling moth, Cydia pomonella (L.), with its sex pheromone (E,E)‐8,10‐dodecadien‐1‐ol (codlemone) alone or in combination with plant volatiles (PH‐plant volatile lures) with and without the addition of an acetic acid (AA) lure was compared in apple and walnut orchards treated with PVC Cidetrak^® dispensers (CM‐DA Combo) loaded with 60 mg pear ester, ethyl (E, Z)‐2,4‐decadienoate (PE) and 185 mg codlemone. Lures were also evaluated in untreated apple and walnut orchards and apple orchards treated with Cidetrak^® dispensers (CM) loaded with 120 mg codlemone. Traps with codlemone lures caught more moths than PE plus AA but similar numbers as traps with the PH‐PE lure in untreated apple and walnut orchards. The PH‐PE outperformed codlemone lures in apple orchards treated with CM dispensers. Codlemone outperformed PH‐PE lures in apple but not in walnut orchards treated mid‐season with CM‐DA Combo dispensers. Combining AA with PE significantly increased moth catch and together they performed similarly to the PH‐PE lure in all dispenser treatments in apple orchards. However, adding AA with the PH‐PE lure did not increase moth catches. The addition of AA to any of the lures tested except codlemone alone significantly increased the catch of females. Adding AA to either PE or the PH‐PE lure created a similarly attractive lure in orchards treated with CM‐DA Combo dispensers. Combining (E)‐4,8‐dimethyl‐1,3,7‐nonatriene (DMNT) with codlemone in a combinational lure plus AA performed as well as any lure tested in apple orchards treated with CM‐DA Combo dispensers. Combinational lures loaded with either (E,E)‐farnesol, (E)‐β‐farnesene or DMNT performed significantly better than the PH‐PE lure in apple orchards treated with CM‐DA Combo dispensers only in the first generation. Both PH‐DMNT and PH‐farnesol lures were comparable to PH‐PE, except that the PH‐farnesol lure was less attractive to female moths.     

Gene editing of the wheat homologs of TONNEAU1‐recruiting motif encoding gene affects grain shape and weight in wheat

Grain size and weight are important components of a suite of yield‐related traits in crops. Here, we showed that the CRISPR‐Cas9 gene editing of TaGW7, a homolog of rice OsGW7 encoding a TONNEAU1‐recruiting motif (TRM) protein, affects grain shape and weight in allohexaploid wheat. By editing the TaGW7 homoeologs in the B and D genomes, we showed that mutations in either of the two or both genomes increased the grain width and weight but reduced the grain length. The effect sizes of mutations in the TaGW7 gene homoeologs coincided with the relative levels of their expression in the B and D genomes. The effects of gene editing on grain morphology and weight traits were dosage dependent with the double‐copy mutant showing larger effect than the respective single copy mutants. The TaGW7‐centered gene co‐expression network indicated that this gene is involved in the pathways regulating cell division and organ growth, also confirmed by the cellular co‐localization of TaGW7 with α‐ and β‐tubulin proteins, the building blocks of microtubule arrays. The analyses of exome capture data in tetraploid domesticated and wild emmer, and hexaploid wheat revealed the loss of diversity around TaGW7‐associated with domestication selection, suggesting that TaGW7 is likely to play an important role in the evolution of yield component traits in wheat. Our study showed how integrating CRISPR‐Cas9 system with cross‐species comparison can help to uncover the function of a gene fixed in wheat for allelic variants targeted by domestication selection and select targets for engineering new gene variants for crop improvement.     

‘Real time’ genetic manipulation: a new tool for ecological field studies

Field experiments with transgenic plants often reveal the functional significance of genetic traits that are important for the performance of the plants in their natural environments. Until now, only constitutive overexpression, ectopic expression and gene silencing methods have been used to analyze gene‐related phenotypes in natural habitats. These methods do not allow sufficient control over gene expression for the study of ecological interactions in real time, of genetic traits that play essential roles in development, or of dose‐dependent effects. We applied the sensitive dexamethasone (DEX)‐inducible pOp6/LhGR expression system to the ecological model plant Nicotiana attenuata and established a lanolin‐based DEX application method to facilitate ectopic gene expression and RNA interference‐mediated gene silencing in the field and under challenging conditions (e.g. high temperature, wind and UV radiation). Fully established field‐grown plants were used to silence phytoene desaturase and thereby cause photobleaching only in specific plant sectors, and to activate expression of the cytokinin (CK) biosynthesis gene isopentenyl transferase (ipt). We used ipt expression to analyze the role of CKs in both the glasshouse and the field to understand resistance to the native herbivore Tupiocoris notatus, which attacks plants at small spatial scales. By spatially restricting ipt expression and elevating CK levels in single leaves, damage by T. notatus increased, demonstrating the role of CKs in this plant–herbivore interaction at a small scale. As the arena of most ecological interactions is highly constrained in time and space, these tools will advance the genetic analysis of dynamic traits that matter for plant performance in nature.     

Haplotype hitchhiking promotes trait coselection in Brassica napus

Local haplotype patterns surrounding densely spaced DNA markers with significant trait associations can reveal information on selective sweeps and genome diversity associated with important crop traits. Relationships between haplotype and phenotype diversity, coupled with analysis of gene content in conserved haplotype blocks, can provide insight into coselection for nonrelated traits. We performed genome‐wide analysis of haplotypes associated with the important physiological and agronomic traits leaf chlorophyll and seed glucosinolate content, respectively, in the major oilseed crop species Brassica napus. A locus on chromosome A01 showed opposite effects on leaf chlorophyll content and seed glucosinolate content, attributed to strong linkage disequilibrium (LD) between orthologues of the chlorophyll biosynthesis genes EARLY LIGHT‐INDUCED PROTEIN and CHLOROPHYLL SYNTHASE, and the glucosinolate synthesis gene ATP SULFURYLASE 1. Another conserved haplotype block, on chromosome A02, contained a number of chlorophyll‐related genes in LD with orthologues of the key glucosinolate biosynthesis genes METHYLTHIOALKYMALATE SYNTHASE‐LIKE 1 and 3. Multigene haplogroups were found to have a significantly greater contribution to variation for chlorophyll content than haplotypes for any single gene, suggesting positive effects of additive locus accumulation. Detailed reanalysis of population substructure revealed a clade of ten related accessions exhibiting high leaf chlorophyll and low seed glucosinolate content. These accessions each carried one of the above‐mentioned haplotypes from A01 or A02, generally in combination with further chlorophyll‐associated haplotypes from chromosomes A05 and/or C05. The phenotypic rather than pleiotropic correlations between leaf chlorophyll content index and seed GSL suggest that LD may have led to inadvertent coselection for these two traits.     

Interaction of acid exudates in chickpea with biological activity of Bacillus thuringiensis towards Helicoverpa armigera

The gram pod borer, Helicoverpa armigera, is one of the most important constraints to chickpea production. High acidity of chickpea exudates is associated with resistance to pod borer, H. armigera; however, acidic exudates in chickpea might influence the biological activity of the bacterium, Bacillus thuringiensis (Bt), applied as a foliar spray or deployed in transgenic plants for controlling H. armigera. Therefore, studies were undertaken to evaluate the biological activity of Bt towards H. armigera on chickpea genotypes with different amounts of organic acids. Significantly lower leaf feeding, larval survival and larval weights were observed on ICC 506EB, followed by C 235, and ICCV 10 across Bt concentrations. Leaf feeding by the larvae and larval survival and weights decreased with an increase in Bt concentration. However, rate of decrease in leaf feeding and larval survival and weights with an increase in Bt concentration was greater on L 550 and ICCV 10 than on the resistant check, ICC 506EB, suggesting that factors in the resistant genotypes, particularly the acid exudates, resulted in lower levels of biological activity of Bt possibly because of antifeedant effects of the acid exudates. Antifeedant effects of acid exudates reduced food consumption and hence might reduce the efficacy of Bt sprays on insect‐resistant chickpea genotypes or Bt‐transgenic chickpeas, although the combined effect of plant resistance based on organic acids, and Bt had a greater effect on survival and development of H. armigera than Bt alone.     

Chromatin interacting factor OsVIL2 increases biomass and rice grain yield

Grain number is an important agronomic trait. We investigated the roles of chromatin interacting factor Oryza sativa VIN3‐LIKE 2 (OsVIL2), which controls plant biomass and yield in rice. Mutations in OsVIL2 led to shorter plants and fewer grains whereas its overexpression (OX) enhanced biomass production and grain numbers when compared with the wild type. RNA‐sequencing analyses revealed that 1958 genes were up‐regulated and 2096 genes were down‐regulated in the region of active division within the first internodes of OX plants. Chromatin immunoprecipitation analysis showed that, among the downregulated genes, OsVIL2 was directly associated with chromatins in the promoter region of CYTOKININ OXIDASE/DEHYDROGENASE2 (OsCKX2), a gene responsible for cytokinin degradation. Likewise, active cytokinin levels were increased in the OX plants. We conclude that OsVIL2 improves the production of biomass and grain by suppressing OsCKX2 chromatin.     

Genetic dissection of photosynthetic efficiency traits for enhancing seed yield in chickpea

Understanding the genetic basis of photosynthetic efficiency (PE) contributing to enhanced seed yield per plant (SYP) is vital for genomics‐assisted crop improvement of chickpea. The current study employed an integrated genomic strategy involving photosynthesis pathway gene‐based association mapping, genome‐wide association study, quantitative trait loci (QTL) mapping, and expression profiling. This identified 16 potential single nucleotide polymorphism loci linked to major QTLs underlying 16 candidate genes significantly associated with PE and SYP traits in chickpea. The allelic variants were tightly linked to positively interacting QTLs regulating both enhanced PE and SYP traits as exemplified by a chlorophyll A‐B binding protein‐coding gene. The leaf tissue‐specific pronounced up‐regulated expression of 16 associated genes in germplasm accessions and homozygous individuals of mapping population was evident. Such combinatorial genomic strategy coupled with gene haplotype‐specific association and in silico protein–protein interaction study delineated natural alleles and superior haplotypes from a chlorophyll A‐B binding (CAB) protein‐coding gene and its interacting gene, Timing of CAB Expression 1 (TOC1), which appear to be most promising candidates in modulating chickpea PE and SYP traits. These functionally pertinent molecular signatures identified have efficacy to drive marker‐assisted selection for developing PE‐enriched cultivars with high seed yield in chickpea.     

Nucleotide variability and haplotype heterogeneity at the porcine fat mass and obesity‐associated (FTO) gene

A large number of studies have confirmed that variants within the fat mass and obesity‐associated (FTO) gene are associated with higher obesity risk in humans. We and others have shown that FTO polymorphisms are associated with fat deposition and related traits in several pig populations, thus confirming the role of this gene in fatness across species. However, some differences observed in different pig populations may be derived, at least in part, from genetic heterogeneity at this locus. Here, we characterise the nucleotide variability and haplotype diversity of the porcine FTO gene in breeds having different predispositions to fat deposition traits. We resequenced 4749 bp of coding and non‐coding regions of the porcine FTO gene in 44 pigs of eight different breeds and identified 27 single nucleotide polymorphisms (SNPs) and four insertions/deletions. A positive Tajima's D‐value (P < 0.10) obtained in Italian Duroc pigs may be compatible with putative balancing selection. From the sequenced pig panel, 20 haplotypes were inferred, some of which clustered according to the breed of origin (Meishan and Italian Duroc). Genetic heterogeneity at this locus could complicate the dissection of the effects of this gene on fat deposition and production traits in pigs. This situation resembles, to some extent, what has been reported in humans, thus making the study of the porcine FTO gene variability especially interesting, as it could be used as a model to understand the complex and elusive role of this gene in human obesity.