The CYP80A and CYP80G Are Involved in the Biosynthesis of Benzylisoquinoline Alkaloids in the Sacred Lotus (Nelumbo nucifera).

Bisbenzylisoquinoline and aporphine alkaloids are the two main pharmacological compounds in the ancient sacred lotus (Nelumbo nucifera). The biosynthesis of bisbenzylisoquinoline and aporphine alkaloids has attracted extensive attention because bisbenzylisoquinoline alkaloids have been reported as potential therapeutic agents for COVID-19. Our study showed that NnCYP80A can catalyze C-O coupling in both (R)-N-methylcoclaurine and (S)-N-methylcoclaurine to produce bisbenzylisoquinoline alkaloids with three different linkages. In addition, NnCYP80G catalyzed C-C coupling in aporphine alkaloids with extensive substrate selectivity, specifically using (R)-N-methylcoclaurine, (S)-N-methylcoclaurine, coclaurine and reticuline as substrates, but the synthesis of C-ring alkaloids without hydroxyl groups in the lotus remains to be elucidated. The key residues of NnCYP80G were also studied using the 3D structure of the protein predicted using Alphafold 2, and six key amino acids (G39, G69, A211, P288, R425 and C427) were identified. The R425A mutation significantly decreased the catalysis of (R)-N-methylcoclaurine and coclaurine inactivation, which might play important role in the biosynthesis of alkaloids with new configurations.

Climate change enhances stability of wheat-flowering-date.

The stability of winter wheat-flowering-date is crucial for ensuring consistent and robust crop performance across diverse climatic conditions. However, the impact of climate change on wheat-flowering-dates remains uncertain. This study aims to elucidate the influence of climate change on wheat-flowering-dates, predict how projected future climate conditions will affect flowering date stability, and identify the most stable wheat genotypes in the study region. We applied a multi-locus genotype-based (MLG-based) model for simulating wheat-flowering-dates, which we calibrated and evaluated using observed data from the Northern China winter wheat region (NCWWR). This MLG-based model was employed to project flowering dates under different climate scenarios. The simulated flowering dates were then used to assess the stability of flowering dates under varying allelic combinations in projected climatic conditions. Our MLG-based model effectively simulated flowering dates, with a root mean square error (RMSE) of 2.3 days, explaining approximately 88.5 % of the genotypic variation in flowering dates among 100 wheat genotypes. We found that, in comparison to the baseline climate, wheat-flowering-dates are expected to shift earlier within the target sowing window by approximately 11 and 14 days by 2050 under the Representative Concentration Pathways 4.5 (RCP4.5) and RCP8.5 climate scenarios, respectively. Furthermore, our analysis revealed that wheat-flowering-date stability is likely to be further strengthened under projected climate scenarios due to early flowering trends. Ultimately, we demonstrate that the combination of Vrn and Ppd genes, rather than individual Vrn or Ppd genes, plays a critical role in wheat-flowering-date stability. Our results suggest that the combination of Ppd-D1a with winter genotypes carrying the vrn-D1 allele significantly contributes to flowering date stability under current and projected climate scenarios. These findings provide valuable insights for wheat breeders and producers under future climatic conditions.

Combined linkage analysis and association mapping identifies genomic regions associated with yield-related and drought-tolerance traits in wheat (Triticum aestivum L.).

KEY MESSAGE: Combined linkage analysis and association mapping identified genomic regions associated with yield and drought tolerance, providing information to assist breeding for high yield and drought tolerance in wheat. Wheat (Triticum aestivum L.) is one of the most widely grown food crops and provides adequate amounts of protein to support human health. Drought stress is the most important abiotic stress constraining yield during the flowering and grain development periods. Precise targeting of genomic regions underlying yield- and drought tolerance-responsive traits would assist in breeding programs. In this study, two water treatments (well-watered, WW, and rain-fed water stress, WS) were applied, and five yield-related agronomic traits (plant height, PH; spike length, SL; spikelet number per spike, SNPS; kernel number per spike, KNPS; thousand kernel weight, TKW) and drought response values (DRVs) were used to characterize the drought sensitivity of each accession. Association mapping was performed on an association panel of 304 accessions, and linkage analysis was applied to a doubled haploid (DH) population of 152 lines. Eleven co-localized genomic regions associated with yield traits and DRV were identified in both populations. Many previously cloned key genes were located in these regions. In particular, a TKW-associated region on chromosome 2D was identified using both association mapping and linkage analysis and a key candidate gene, TraesCS2D02G142500, was detected based on gene annotation and differences in expression levels. Exonic SNPs were analyzed by sequencing the full length of TraesCS2D02G142500 in the association panel, and a rare haplotype, Hap-2, which reduced TKW to a lesser extent than Hap-1 under drought stress, and the Hap-2 varieties presented drought-insensitive. Altogether, this study provides fundamental insights into molecular targets for high yield and drought tolerance in wheat.

A high-resolution genotype-phenotype map identifies the TaSPL17 controlling grain number and size in wheat.

BACKGROUND: Large-scale genotype-phenotype association studies of crop germplasm are important for identifying alleles associated with favorable traits. The limited number of single-nucleotide polymorphisms (SNPs) in most wheat genome-wide association studies (GWASs) restricts their power to detect marker-trait associations. Additionally, only a few genes regulating grain number per spikelet have been reported due to sensitivity of this trait to variable environments. RESULTS: We perform a large-scale GWAS using approximately 40 million filtered SNPs for 27 spike morphology traits. We detect 132,086 significant marker-trait associations and the associated SNP markers are located within 590 associated peaks. We detect additional and stronger peaks by dividing spike morphology into sub-traits relative to GWAS results of spike morphology traits. We propose that the genetic dissection of spike morphology is a powerful strategy to detect signals for grain yield traits in wheat. The GWAS results reveal that TaSPL17 positively controls grain size and number by regulating spikelet and floret meristem development, which in turn leads to enhanced grain yield per plant. The haplotypes at TaSPL17 indicate geographical differentiation, domestication effects, and breeding selection. CONCLUSION: Our study provides valuable resources for genetic improvement of spike morphology and a fast-forward genetic solution for candidate gene detection and cloning in wheat.

TaTIP41 and TaTAP46 positively regulate drought tolerance in wheat by inhibiting PP2A activity.

Drought is a major environmental stress limiting global wheat (Triticum aestivum) production. Exploring drought tolerance genes is important for improving drought adaptation in this crop. Here, we cloned and characterized TaTIP41, a novel drought tolerance gene in wheat. TaTIP41 is a putative conserved component of target of rapamycin (TOR) signaling, and the TaTIP41 homoeologs were expressed in response to drought stress and abscisic acid (ABA). The overexpression of TaTIP41 enhanced drought tolerance and the ABA response, including ABA-induced stomatal closure, while its downregulation using RNA interference (RNAi) had the opposite effect. Furthermore, TaTIP41 physically interacted with TaTAP46, another conserved component of TOR signaling. Like TaTIP41, TaTAP46 positively regulated drought tolerance. Furthermore, TaTIP41 and TaTAP46 interacted with type-2A protein phosphatase (PP2A) catalytic subunits, such as TaPP2A-2, and inhibited their enzymatic activities. Silencing TaPP2A-2 improved drought tolerance in wheat. Together, our findings provide new insights into the roles of TaTIP41 and TaTAP46 in the drought tolerance and ABA response in wheat, and their potential application in improving wheat environmental adaptability.

Wheat NAC-A18 regulates grain starch and storage proteins synthesis and affects grain weight.

KEY MESSAGE: Wheat NAC-A18 regulates both starch and storage protein synthesis in the grain, and a haplotype with positive effects on grain weight showed increased frequency during wheat breeding in China. Starch and seed storage protein (SSP) directly affect the processing quality of wheat grain. The synthesis of starch and SSP are also regulated at the transcriptional level. However, only a few starch and SSP regulators have been identified in wheat. In this study, we discovered a NAC transcription factor, designated as NAC-A18, which acts as a regulator of both starch and SSP synthesis. NAC-A18, is predominately expressed in wheat developing grains, encodes a transcription factor localized in the nucleus, with both activation and repression domains. Ectopic expression of wheat NAC-A18 in rice significantly decreased starch accumulation and increased SSP accumulation and grain size and weight. Dual-luciferase reporter assays indicated that NAC-A18 could reduce the expression of TaGBSSI-A1 and TaGBSSI-A2, and enhance the expression of TaLMW-D6 and TaLMW-D1. A yeast one hybrid assay demonstrated that NAC-A18 bound directly to the cis-element "ACGCAA" in the promoters of TaLMW-D6 and TaLMW-D1. Further analysis indicated that two haplotypes were formed at NAC-A18, and that NAC-A18_h1 was a favorable haplotype correlated with higher thousand grain weight. Based on limited population data, NAC-A18_h1 underwent positive selection during Chinese wheat breeding. Our study demonstrates that wheat NAC-A18 regulates starch and SSP accumulation and grain size. A molecular marker was developed for the favorable allele for breeding applications.

Genetic basis of geographical differentiation and breeding selection for wheat plant architecture traits.

BACKGROUND: Plant architecture associated with increased grain yield and adaptation to the local environments is selected during wheat (Triticum aestivum) breeding. The internode length of individual stems and tiller length of individual plants are important for the determination of plant architecture. However, few studies have explored the genetic basis of these traits. RESULTS: Here, we conduct a genome-wide association study (GWAS) to dissect the genetic basis of geographical differentiation of these traits in 306 worldwide wheat accessions including both landraces and traditional varieties. We determine the changes of haplotypes for the associated genomic regions in frequency in 831 wheat accessions that are either introduced from other countries or developed in China from last two decades. We identify 83 loci that are associated with one trait, while the remaining 247 loci are pleiotropic. We also find 163 associated loci are under strong selective sweep. GWAS results demonstrate independent regulation of internode length of individual stems and consistent regulation of tiller length of individual plants. This makes it possible to obtain ideal haplotype combinations of the length of four internodes. We also find that the geographical distribution of the haplotypes explains the observed differences in internode length among the worldwide wheat accessions. CONCLUSION: This study provides insights into the genetic basis of plant architecture. It will facilitate gene functional analysis and molecular design of plant architecture for breeding.

Visualization and identification of benzylisoquinoline alkaloids in various nelumbo nucifera tissues.

Benzylisoquinoline alkaloids in lotus (Nelumbo nucifera) seed plumules and leaves exhibit significant tissue specificity for their pharmacological effects and potential nutritional properties. Herein, 46 benzylisoquinoline alkaloids were identified via UPLC-QTOF-HRMS, of which 9 were annotated as glycosylated monobenzylisoquinoline alkaloids concentrated in the seed plumules. The spatial distribution of targeted benzylisoquinoline alkaloids in leaves, seed plumules, and milky sap was determined via MALDI-MSI. Furthermore, 37 Nelumbo cultivars were investigated using targeted metabolomics to provide insights into functional tea development. While aporphine alkaloids comprised the main compounds present in lotus leaves, bisbenzylisoquinoline alkaloids were the main compounds in lotus plumules, where glycosylation primarily occurs. These findings can help understand the distribution of benzylisoquinoline alkaloids in lotus tissue and the directional breeding of varieties enriched with specific chemical functional groups for nutritional and pharmacological applications.

TaSPL14-7A is a conserved regulator controlling plant architecture and yield traits in common wheat (Triticum aestivum L.).

Plant architecture is a crucial influencing factor of wheat yield and adaptation. In this study, we cloned and characterized TaSPL14, a homologous gene of the rice ideal plant architecture gene OsSPL14 in wheat. TaSPL14 homoeologs (TaSPL14-7A, TaSPL14-7B and TaSPL14-7D) exhibited similar expression patterns, and they were all preferentially expressed in stems at the elongation stage and in young spikes. Moreover, the expression level of TaSPL14-7A was higher than that of TaSPL14-7B and TaSPL14-7D. Overexpression of TaSPL14-7A in wheat resulted in significant changes in plant architecture and yield traits, including decreased tiller number and increased kernel size and weight. Three TaSPL14-7A haplotypes were identified in Chinese wheat core collection, and haplotype-based association analysis showed that TaSPL14-7A-Hap1/2 were significantly correlated with fewer tillers, larger kernels and higher kernel weights in modern cultivars. The haplotype effect resulted from a difference in TaSPL14-7A expression levels among genotypes, with TaSPL14-7A-Hap1/2 leading to higher expression levels than TaSPL14-7A-Hap3. As favorable haplotypes, TaSPL14-7A-Hap1/2 underwent positive selection during global wheat breeding over the last century. Together, the findings of our study provide insight into the function and genetic effects of TaSPL14 and provide a useful molecular marker for wheat breeding.

TabHLH95-TaNF-YB1 module promotes grain starch synthesis in bread wheat.

Starch is the most abundant substance in wheat (Triticum aestivum L.) endosperm and provides the major carbohydrate energy for human daily life. Starch synthesis-related (SSR) genes are believed to be spatiotemporally specific, but their transcriptional regulation remains unclear in wheat. Here, we investigate the role of the basic helix-loop-helix (bHLH) transcription factor TabHLH95 in starch synthesis. TabHLH95 is preferentially expressed in the developing grains in wheat and encodes a nucleus localized protein without autoactivation activity. The Tabhlh95 knockout mutants display smaller grain size and less starch content than wild type, whereas overexpression of TabHLH95 enhances starch accumulation and significantly improves thousand grain weight. Transcriptome analysis reveals that the expression of multiple SSR genes is significantly reduced in the Tabhlh95 mutants. TabHLH95 binds to the promoters of ADP-glucose pyrophosphorylase large subunit 1 (AGPL1-1D/-1B), AGPL2-5D, and isoamylase (ISA1-7D) and enhances their transcription. Intriguingly, TabHLH95 interacts with the nuclear factor Y (NF-Y) family transcription factor TaNF-YB1, thereby synergistically regulating starch synthesis. These results suggest that the TabHLH95-TaNF-YB1 complex positively modulates starch synthesis and grain weight by regulating the expression of a subset of SSR genes, thus providing a good potential approach for genetic improvement of grain productivity in wheat.

Transcriptome Analysis of Roots from Wheat (Triticum aestivum L.) Varieties in Response to Drought Stress.

Under climate change, drought is one of the most limiting factors that influences wheat (Triticum aestivum L.) production. Exploring stress-related genes is vital for wheat breeding. To identify genes related to the drought tolerance response, two common wheat cultivars, Zhengmai 366 (ZM366) and Chuanmai 42 (CM42), were selected based on their obvious difference in root length under 15% PEG-6000 treatment. The root length of the ZM366 cultivar was significantly longer than that of CM42. Stress-related genes were identified by RNA-seq in samples treated with 15% PEG-6000 for 7 days. In total, 11,083 differentially expressed genes (DEGs) and numerous single nucleotide polymorphisms (SNPs) and insertions/deletions (InDels) were identified. GO enrichment analysis revealed that the upregulated genes were mainly related to the response to water, acidic chemicals, oxygen-containing compounds, inorganic substances, and abiotic stimuli. Among the DEGs, the expression levels of 16 genes in ZM366 were higher than those in CM42 after the 15% PEG-6000 treatment based on RT-qPCR. Furthermore, EMS-induced mutants in Kronos (T. turgidum L.) of 4 representative DEGs possessed longer roots than the WT after the 15% PEG-6000 treatment. Altogether, the drought stress genes identified in this study represent useful gene resources for wheat breeding.

HSP90.2 promotes CO2 assimilation rate, grain weight and yield in wheat.

Wheat fixes CO2 by photosynthesis into kernels to nourish humankind. Improving the photosynthesis rate is a major driving force in assimilating atmospheric CO2 and guaranteeing food supply for human beings. Strategies for achieving the above goal need to be improved. Here, we report the cloning and mechanism of CO2 ASSIMILATION RATE AND KERNEL-ENHANCED 1 (CAKE1) from durum wheat (Triticum turgidum L. var. durum). The cake1 mutant displayed a lower photosynthesis rate with smaller grains. Genetic studies identified CAKE1 as HSP90.2-B, encoding cytosolic molecular chaperone folding nascent preproteins. The disturbance of HSP90.2 decreased leaf photosynthesis rate, kernel weight (KW) and yield. Nevertheless, HSP90.2 over-expression increased KW. HSP90.2 recruited and was essential for the chloroplast localization of nuclear-encoded photosynthesis units, for example PsbO. Actin microfilaments docked on the chloroplast surface interacted with HSP90.2 as a subcellular track towards chloroplasts. A natural variation in the hexaploid wheat HSP90.2-B promoter increased its transcription activity, enhanced photosynthesis rate and improved KW and yield. Our study illustrated an HSP90.2-Actin complex sorting client preproteins towards chloroplasts to promote CO2 assimilation and crop production. The beneficial haplotype of Hsp90.2 is rare in modern varieties and could be an excellent molecular switch promoting photosynthesis rate to increase yield in future elite wheat varieties.

TaTPP-7A positively feedback regulates grain filling and wheat grain yield through T6P-SnRK1 signalling pathway and sugar-ABA interaction.

Grain size and filling are two key determinants of grain thousand-kernel weight (TKW) and crop yield, therefore they have undergone strong selection since cereal was domesticated. Genetic dissection of the two traits will improve yield potential in crops. A quantitative trait locus significantly associated with wheat grain TKW was detected on chromosome 7AS flanked by a simple sequence repeat marker of Wmc17 in Chinese wheat 262 mini-core collection by genome-wide association study. Combined with the bulked segregant RNA-sequencing (BSR-seq) analysis of an F2 genetic segregation population with extremely different TKW traits, a candidate trehalose-6-phosphate phosphatase gene located at 135.0 Mb (CS V1.0), designated as TaTPP-7A, was identified. This gene was specifically expressed in developing grains and strongly influenced grain filling and size. Overexpression (OE) of TaTPP-7A in wheat enhanced grain TKW and wheat yield greatly. Detailed analysis revealed that OE of TaTPP-7A significantly increased the expression levels of starch synthesis- and senescence-related genes involved in abscisic acid (ABA) and ethylene pathways. Moreover, most of the sucrose metabolism and starch regulation-related genes were potentially regulated by SnRK1. In addition, TaTPP-7A is a crucial domestication- and breeding-targeted gene and it feedback regulates sucrose lysis, flux, and utilization in the grain endosperm mainly through the T6P-SnRK1 pathway and sugar-ABA interaction. Thus, we confirmed the T6P signalling pathway as the central regulatory system for sucrose allocation and source-sink interactions in wheat grains and propose that the trehalose pathway components have great potential to increase yields in cereal crops.

Conservatively transmitted alleles of key agronomic genes provide insights into the genetic basis of founder parents in bread wheat (Triticum aestivum L.).

BACKGROUND: Founder parents play extremely important roles in wheat breeding. Studies into the genetic basis of founder parents and the transmission rules of favorable alleles are of great significance in improving agronomically important traits in wheat. RESULTS: Here, a total of 366 founder parents, widely grown cultivars, and derivatives of four representative founder parents were genotyped based on efficient kompetitive allele-specific PCR (KASP) markers in 87 agronomically important genes controlling yield, quality, adaptability, and stress resistance. Genetic composition analysis of founder parents and widely grown cultivars showed a consistently high frequency of favorable alleles for yield-related genes. This analysis further showed that other alleles favorable for resistance, strong gluten, dwarf size, and early heading date were also subject to selective pressure over time. By comparing the transmission of alleles from four representative founder parents to their derivatives during different breeding periods, it was found that the genetic composition of the representative founder parents was optimized as breeding progressed over time, with the number and types of favorable alleles carried gradually increasing and becoming enriched. There are still a large number of favorable alleles in wheat founder parents that have not been fully utilized in breeding selection. Eighty-seven agronomically important genes were used to construct an enrichment map that shows favorable alleles of four founder parents, providing an important theoretical foundation for future identification of candidate wheat founder parents. CONCLUSIONS: These results reveal the genetic basis of founder parents and allele transmission for 87 agronomically important genes and shed light on breeding strategies for the next generation of elite founder parents in wheat.

Fast integration and accumulation of beneficial breeding alleles through an AB-NAMIC strategy in wheat.

Wheat (Triticum aestivum) is among the most important staple crops for safeguarding the food security of the growing world population. To bridge the gap between genebank diversity and breeding programs, we developed an advanced backcross-nested association mapping plus inter-crossed population (AB-NAMIC) by crossing three popular wheat cultivars as recurrent founders to 20 germplasm lines from a mini core collection. Selective backcrossing combined with selection against undesirable traits and extensive crossing within and between sub-populations created new opportunities to detect unknown genes and increase the frequency of beneficial alleles in the AB-NAMIC population. We performed phenotyping of 590 AB-NAMIC lines and a natural panel of 476 cultivars for six consecutive growing seasons and genotyped these 1066 lines with a 660K SNP array. Genome-wide association studies of both panels for plant development and yield traits demonstrated improved power to detect rare alleles and loci with medium genetic effects in AB-NAMIC. Notably, genome-wide association studies in AB-NAMIC detected the candidate gene TaSWEET6-7B (TraesCS7B03G1216700), which has high homology to the rice SWEET6b gene and exerts strong effects on adaptation and yield traits. The commercial release of two derived AB-NAMIC lines attests to its direct applicability in wheat improvement. Valuable information on genome-wide association study mapping, candidate genes, and their haplotypes for breeding traits are available through WheatGAB. Our research provides an excellent framework for fast-tracking exploration and accumulation of beneficial alleles stored in genebanks.

The auxin response factor TaARF15-A1 negatively regulates senescence in common wheat (Triticum aestivum L.).

Auxin plays an important role in regulating leaf senescence. Auxin response factors (ARFs) are crucial components of the auxin signaling pathway; however, their roles in leaf senescence in cereal crops are unknown. In this study, we identified TaARF15-A1 as a negative regulator of senescence in wheat (Triticum aestivum L.) by analyzing TaARF15-A1 overexpression (OE) and RNA interference lines and CRISPR/Cas9-based arf15 mutants. OE of TaARF15-A1 delayed senescence, whereas knockdown lines and knockout mutants showed accelerated leaf senescence and grain ripening. RNA-seq analysis revealed that TaARF15-A1 delays leaf senescence by negatively regulating senescence-promoting processes and positively modulating senescence-delaying genes including senescence-associated phytohormone biosynthesis and metabolism genes as well as transcription factors (TFs). We also demonstrated that TaARF15-A1 physically interacts with TaMYC2, a core jasmonic acid (JA) signaling TF that positively modulates wheat senescence. Furthermore, TaARF15-A1 suppressed the expression of TaNAM-1 (TaNAM-A1 and TaNAM-D1) via protein-protein interaction and competition with TaMYC2 for binding to its promoter to regulate senescence. Finally, we identified two haplotypes of TaARF15-A1 in global wheat collections. Association analysis revealed that TaARF15-A1-HapI has undergone strong selection during wheat breeding in China, likely owing to its earlier maturity. Thus, we identify TaARF15-A1 as a negative regulator of senescence in common wheat and present another perspective on the crosstalk between auxin and JA signaling pathways in regulating plant senescence.

Identification of the major QTL QPm.cas-7D for adult plant resistance to wheat powdery mildew.

Developing effective and durable host plant resistance is crucial for controlling powdery mildew, a devastating disease caused by Blumeria graminis f. sp. tritici (Bgt). In the present study, we dissected the genetic basis of the adult plant resistance to powdery mildew using a recombinant inbred line (RIL) composed of 176 F9 RILs population derived from a cross between PuBing 3228 (P3228) and susceptible cultivar Gao 8901. P3228 exhibits stable adult-plant resistance to powdery mildew in the field over consecutive years. We identified two QTLs on chromosomes 7DS (QPm.cas-7D) and 1AL (QPm.cas-1A) contributed by P3228, and one QTL on 3DS (QPm.cas-3D) contributed by Gao 8901, which could explain 65.44%, 3.45%, and 2.18% of the phenotypic variances, respectively. By analyzing the annotated genes in the 1.168 Mb physical interval of the major QTL QPm.cas-7D, we locked a previously cloned adult-plant resistance gene Pm38 that was most probably the candidate gene of QPm.cas-7D. Sequence alignment analysis revealed that the candidate gene of QPm.cas-7D in P3228 was identical to the reported Pm38 sequence. Two haplotypes QPm-7D-R and QPm-7D-S were identified in the whole Pm38 genomic regions between P3228 and Gao 8901. To apply QPm.cas-7D in wheat breeding, we developed a kompetitive allele-specific PCR (KASP) marker Kasp5249 that is closely linked with these haplotypes. It is worth mentioning that the QPm-7D-R haplotype significantly decreased TKW and underwent negative selection for higher yields in China wheat breeding. In this study, we identified a major QTL QPm.cas-7D and revealed the relationship between its resistance and yield, which could be beneficial for further applications in wheat disease resistance and high-yield breeding.

TaNAC020 homoeologous genes are associated with higher thousand kernel weight and kernel length in Chinese wheat.

NAC proteins constitute one of the largest plant-specific transcription factor (TF) families and play significant roles in plant growth and development. In the present study, three TaNAC020 homoeologous genes located on chromosomes 7A, 7B, and 7D were isolated from wheat (Triticum aestivum L.). TaNAC020s were predominantly expressed in developing grains. The developed transgenic rice lines for TaNAC020-B showed higher starch density and lower amylose contents than those of the wild type (WT). Sequence polymorphism studies showed seven and eight SNPs in TaNAC020-A/B, making three and two haplotypes, respectively. No sequence polymorphism was identified in TaNAC020-D. Association analysis revealed that HAP-2 of TaNAC020-A and TaNAC020-B was the favored haplotype for higher thousand kernel weight and length. Geographic distribution and allelic frequency showed that our favored haplotype experienced strong selection in China, and likewise, diversity increased in TaNAC020s during wheat polyploidization. The results obtained in this study demonstrate that TaNAC020s positively influence starch synthesis and accumulation and are one of the key regulators of the kernel (seed) size and kernel number and have the potential for utilization in wheat breeding to improve grain yield. Molecular markers developed in this study stand instrumental in marker-assisted selection for genetic improvement and germplasm enhancement in wheat.

Genetic dissection of lutein content in common wheat via association and linkage mapping.

KEY MESSAGE: Genetic architecture controlling grain lutein content of common wheat was investigated through an integration of genome-wide association study (GWAS) and linkage analysis. Putative candidate genes involved in carotenoid metabolism and regulation were identified, which provide a basis for gene cloning and development of nutrient-enriched wheat varieties through molecular breeding. Lutein, known as 'the eye vitamin', is an important component of wheat nutritional and end-use quality. However, the genetic manipulation of grain lutein content (LUC) in common wheat has not previously been well studied. Here, quantitative trait loci (QTL) associated with the LUC measured by high performance liquid chromatography (HPLC) were first identified by integrating a genome-wide association study (GWAS) and linkage mapping. A Chinese wheat mini-core collection (MCC) of 262 accessions and a doubled haploid (DH) population derived from Jinchun 7 and L1219 were genotyped using the 90K SNP array. A total of 124 significant marker-trait associations (MTAs) on all 21 wheat chromosomes except for 1A, 4D, and 5B that formed 58 QTL were detected. Among them, six stable QTL were identified on chromosomes 2AL, 2DS, 3BL, 3DL, 7AL, and 7BS. Meanwhile, three of the ten QTL identified in the DH population, QLuc.5A.1 and QLuc.5A.2 on chromosome 5AL and QLuc.6A.2 on 6AS, were stable and independently explained 5.58-10.86% of the phenotypic variation. The QLuc.6A.2 region colocalized with two MTAs identified by GWAS. Moreover, 71 carotenoid metabolism-related candidate genes were identified, and the allelic effects were analyzed in the MCC panel based on the 90K array. Results revealed that the genes CYP97A3 (Chr. 6B) and CCD1 (Chr. 5A) were significantly associated with LUC. Additionally, the gene PSY3 (QLuc.5A.1) and several candidate genes involved in the methylerythritol 4-phosphate (MEP) pathways colocalized with stable QTL regions. The present study provides potential targets for future functional gene exploration and molecular breeding in common wheat.