TaFlo2-A1, an ortholog of rice Flo2, is associated with thousand grain weight in bread wheat (Triticum aestivum L.).
ABSTRACT: The Flo2 gene is a member of a conserved gene family in plants. This gene has been found to be related to thousand grain weight (TGW) in rice. Its orthologs in hexaploid wheat were cloned, and the haplotype variation in TaFlo2-A1 was tested for association with TGW.The cloned sequences of TaFlo2-A1, TaFlo2-B1 and TaFlo2-D1 contained 23, 23 and 24 exons, respectively. The deduced proteins of TaFlo2-A1 (1734 aa), TaFlo2-B1 (1698 aa) and TaFlo2-D1 (1682 aa) were highly similar (>94%) and exhibited >77% similarity with the rice FLO2 protein. Like the rice FLO2 protein, four tetratricopeptide repeat (TPR) motifs were observed in the deduced TaFLO2 protein. An 8-bp InDel (-10 to -17 bp) in the promoter region and five SNPs in first intron of TaFlo2-A1 together formed two haplotypes, TaFlo2-A1a and TaFlo2-A1b, in bread wheat. TaFlo2 was located on homeologous group 2 chromosomes. TaFlo2-A1 was inferred to be located on deletion bin '2AL1-0.85-1.00'. The TaFlo2-A1 haplotypes were characterized in the Chinese Micro Core Collection (MCC) and Pakistani wheat collection using the molecular marker TaFlo2-Indel8. TaFlo2-A1 was found to be associated with TGW but not with grain number per spike (GpS) in both the MCC and Pakistani wheat collections. The frequency of TaFlo2-A1b (positive haplotype) was low in commercial wheat cultivars; thus this haplotype can be selected to improve grain weight without negatively affecting GpS. The expression level of TaFlo2-A1 in developing grains at 5 DAF (days after flowering) was positively correlated with TGW in cultivars carrying the positive haplotype.This study will likely lead to additional investigations to understand the regulatory mechanism of the Flo2 gene in hexaploid wheat. Furthermore, the newly developed molecular marker 'TaFlo2-InDel8' could be incorporated into the kit of wheat breeders for use in marker-assisted selection.
Project description:Grain yield in wheat is a polygenic trait that is influenced by environmental and genetic interactions at all stages of the plant's growth. Yield is usually broken down into three components; number of spikes per area, grain number per spike, and grain weight (TGW). In polyploid wheat, studies have identified quantitative trait loci (QTL) which affect TGW, yet few have been validated and fine-mapped using independent germplasm, thereby having limited impact in breeding.In this study we identified a major QTL for TGW, yield and green canopy duration on wheat chromosome 6A of the Spark x Rialto population, across 12 North European environments. Using independent germplasm in the form of BC2 and BC4 near isogenic lines (NILs), we validated the three QTL effects across environments. In four of the five experiments the Rialto 6A introgression gave significant improvements in yield (5.5%) and TGW (5.1%), with morphometric measurements showing that the increased grain weight was a result of wider grains. The extended green canopy duration associated with the high yielding/TGW Rialto allele was comprised of two independent effects; earlier flowering and delayed final maturity, and was expressed stably across the five environments. The wheat homologue (TaGW2) of a rice gene associated with increased TGW and grain width was mapped within the QTL interval. However, no polymorphisms were identified in the coding sequence between the parents.The discovery and validation through near-isogenic lines of robust QTL which affect yield, green canopy duration, thousand grain weight, and grain width on chromosome 6A of hexaploid wheat provide an important first step to advance our understanding of the genetic mechanisms regulating the complex processes governing grain size and yield in polyploid wheat.
Project description:In wheat, the vernalization requirement is mainly controlled by the VRN genes. Different species of hexaploid and tetraploid wheat are widely used as genetic source for new mutant variants and alleles for fundamental investigations and practical breeding programs. In this study, VRN-A1 and VRN-B1 were analysed for 178 accessions representing six tetraploid wheat species (Triticum dicoccoides, T. dicoccum, T. turgidum, T. polonicum, T. carthlicum, T. durum) and five hexaploid species (T. compactum, T. sphaerococcum, T. spelta, T. macha, T. vavilovii).Novel allelic variants in the promoter region of VRN-A1 and VRN-B1 were identified based on the change in curvature and flexibility of the DNA molecules. The new variants of VRN-A1 (designated as Vrn-A1a.2, Vrn-A1b.2 - Vrn-A1b.6 and Vrn-A1i) were found to be widely distributed in hexaploid and tetraploid wheat, and in fact were predominant over the known VRN-A1 alleles. The greatest diversity of the new variants of VRN-B1 (designated as VRN-B1.f, VRN-B1.s and VRN-B1.m) was found in the tetraploid and some hexaploid wheat species. For the first time, minor differences within the sequence motif known as the VRN-box of VRN1 were correlated with wheat growth habit. Thus, vrn-A1b.3 and vrn-A1b.4 were revealed in winter wheat in contrast to Vrn-A1b.2, Vrn-A1b.5, Vrn-A1b.6 and Vrn-A1i. It was found that single nucleotide mutation in the VRN-box can influence the vernalization requirement and growth habit of wheat. Our data suggest that both the A-tract and C-rich segment within the VRN-box contribute to its functionality, and provide a new view of the hypothesised role of the VRN-box in regulating transcription of the VRN1 genes. Specifically, it is proposed that combination of mutations in this region can modulate vernalization sensitivity and flowering time of wheat.New allelic variants of the VRN-A1 and VRN-B1 genes were identified in hexaploid and tetraploid wheat. Mutations in A-tract and C-rich segments within the VRN-box of VRN-A1 are associated with modulation of the vernalization requirement and flowering time. New allelic variants will be useful in fundamental investigations into the regulation of VRN1 expression, and provide a valuable genetic resource for practical breeding of wheat.
Project description:Across 13 experiments the gw2 - A1 mutant allele shifts grain size distribution consistently across all grains significantly increasing grain weight (6.6 %), width (2.8 %) and length (2.1 %) in tetraploid and hexaploid wheat. There is an urgent need to identify, understand and incorporate alleles that benefit yield in polyploid wheat. The rice OsGW2 gene functions as a negative regulator of grain weight and width and is homologous to the wheat TaGW2 gene. Previously it was shown that transcript levels of the A-genome homoeologue, TaGW2-A1, are negatively associated with grain width in hexaploid wheat. In this study we screened the tetraploid Kronos TILLING population to identify mutants in TaGW2-A1. We identified a G to A transition in the splice acceptor site of exon 5 which leads to mis-splicing in TaGW2-A1. We backcrossed the mutant allele into tetraploid and hexaploid wheat and generated a series of backcross derived isogenic lines which were evaluated in glasshouse and field conditions. Across 13 experiments the GW2-A1 mutant allele significantly increased thousand grain weight (6.6 %), grain width (2.8 %) and grain length (2.1 %) in tetraploid and hexaploid wheat compared to the wild type allele. In hexaploid wheat, this led to an increase in spike yield since no differences were detected for spikelet or grain number between isogenic lines. The increase in grain width and length was consistent across grains of different sizes, suggesting that the effect of the mutation is stable across the ear and within spikelets. Differences in carpel size and weight between alleles were identified as early as 5 days before anthesis, suggesting that TaGW2-A1 acts on maternal tissue before anthesis to restrict seed size. A single nucleotide polymorphism marker was developed to aid the deployment of the mutant allele into breeding programmes.
Project description:A novel TaGW2-A1 allele was identified from a stable, robust QTL region, which is pleiotropic for thousand grain weight, grain number per spike, and grain morphometric parameters in wheat. Thousand grain weight (TGW) and grain number per spike (GNS) are two crucial determinants of wheat spike yield, and genetic dissection of their relationships can help to fine-tune these two components and maximize grain yield. By evaluating 191 recombinant inbred lines in 11 field trials, we identified five genomic regions on chromosomes 1B, 3A, 3B, 5B, or 7A that solely influenced either TGW or GNS, and a further region on chromosome 6A that concurrently affected TGW and GNS. The QTL of interest on chromosome 6A, which was flanked by wsnp_BE490604A_Ta_2_1 and wsnp_RFL_Contig1340_448996 and designated as QTgw/Gns.cau-6A, was finely mapped to a genetic interval shorter than 0.538 cM using near isogenic lines (NILs). The elite NILs of QTgw/Gns.cau-6A increased TGW by 8.33%, but decreased GNS by 3.05% in six field trials. Grain Weight 2 (TaGW2-A1), a well-characterized gene that negatively regulates TGW and grain width in wheat, was located within the finely mapped interval of QTgw/Gns.cau-6A. A novel and rare TaGW2-A1 allele with a 114-bp deletion in the 5' flanking region was identified in the parent with higher TGW, and it reduced TaGW2-A1 promoter activity and expression. In conclusion, these results expand our knowledge of the genetic and molecular basis of TGW-GNS trade-offs in wheat. The QTLs and the novel TaGW2-A1 allele are likely useful for the development of cultivars with higher TGW and/or higher GNS.
Project description:TaGW2 is an orthologue of rice gene OsGW2, which encodes E3 RING ubiquitin ligase and controls the grain size in rice. In wheat, three copies of TaGW2 have been identified and mapped on wheat homoeologous group 6 viz. TaGW2-6A, TaGW2-6B and TaGW2-6D. In the present study, using as many as 207 Indian wheat genotypes, we identified four SNPs including two novel SNPs (SNP-988 and SNP-494) in the promoter sequence of TaGW2-6A. All the four SNPs were G/A or A/G substitutions (transitions). Out of the four SNPs, SNP-494 was causal, since it was found associated with grain weight. The mean TGW (41.1 g) of genotypes with the allele SNP-494_A was significantly higher than mean TGW (38.6 g) of genotypes with the allele SNP-494_G. SNP-494 also regulates the expression of TaGW2-6A so that the wheat genotypes with SNP-494_G have higher expression and lower TGW and the genotypes with SNP-494_A have lower expression but higher TGW. Besides, SNP-494 was also found associated with grain length-width ratio, awn length, spike length, grain protein content, peduncle length and plant height. This suggested that gene TaGW2-6A not only controls grain size, but also controls other agronomic traits. In the promoter region, SNP-494 was present in 'CGCG' motif that plays an important role in Ca2+/calmodulin mediated regulation of genes. A user-friendly CAPS marker was also developed to identify the desirable allele of causal SNP (SNP-494) for use in marker-assisted selection for improvement of grain weight in wheat. Using four SNPs, five haplotypes were identified; of these, Hap_5 (G_A_G_A) was found to be a desirable haplotype having significantly higher grain weight (41.13g) relative to other four haplotypes (36.33-39.16 g).
Project description:Increased thousand-grain weight (TGW) is an important breeding target for indirectly improving grain yield (GY). Fourteen reported candidate genes known to enhance TGW were evaluated in two independent and existing datasets of wheat at CIMMYT, the Elite Yield Trial (EYT) from 2015 to 2016 (EYT2015-16) and the Wheat Association Mapping Initiative (WAMI) panel, to study their allele effects on TGW and to verify their suitability for marker-assisted selection. Of these, significant associations were detected for only one gene (TaGs3-D1) in the EYT2015-16 and two genes (TaTGW6 and TaSus1) in WAMI. The reported favorable alleles of TaGs3-D1 and TaTGW6 genes decreased TGW in the datasets. A haplotype-based genome wide association study was implemented to identify the genetic determinants of TGW on a large set of CIMMYT germplasm (4,302 lines comprising five EYTs), which identified 15 haplotype blocks to be significantly associated with TGW. Four of them, identified on chromosomes 4A, 6A, and 7A, were associated with TGW in at least three EYTs. The locus on chromosome 6A (Hap-6A-13) had the largest effect on TGW and additionally GY with increases of up to 2.60 g and 258 kg/ha, respectively. Discovery of novel TGW loci described in our study expands the opportunities for developing diagnostic markers and for multi-gene pyramiding to derive new allele combinations for enhanced TGW and GY in CIMMYT wheat.
Project description:Grain weight, an essential yield component, is under strong genetic control and markedly influenced by the environment. Here, by genome-wide association analysis with a panel of 94 elite common wheat varieties, 37 loci were found significantly associated with thousand-grain weight (TGW) in one or more environments differing in water and fertiliser levels. Five loci were stably associated with TGW under all 12 environments examined. Their elite alleles had positive effects on TGW. Four, two, three, and two loci were consistently associated with TGW in the irrigated and fertilised (IF), rainfed (RF), reduced nitrogen (RN), and reduced phosphorus (RP) environments. The elite alleles of the IF-specific loci enhanced TGW under well-resourced conditions, whereas those of the RF-, RN-, or RP-specific loci conferred tolerance to the TGW decrease when irrigation, nitrogen, or phosphorus were reduced. Moreover, the elite alleles of the environment-independent and -specific loci often acted additively to enhance TGW. Four additional loci were found associated with TGW in specific locations, one of which was shown to contribute to the TGW difference between two experimental sites. Further analysis of 14 associated loci revealed that nine affected both grain length and width, whereas the remaining loci influenced either grain length or width, indicating that these loci control grain weight by regulating kernel size. Finally, the elite allele of Xpsp3152 frequently co-segregated with the larger grain haplotype of TaGW2-6A, suggesting probable genetic and functional linkages between Xpsp3152 and GW2 that are important for grain weight control in cereal plants. Our study provides new knowledge on TGW control in elite common wheat lines, which may aid the improvement of wheat grain weight trait in further research.
Project description:Modern breeding imposed selection for improved productivity that largely influenced the frequency of superior alleles underpinning traits of breeding interest. Therefore, molecular diagnosis for the allelic variations of such genes is important to manipulate beneficial alleles in wheat molecular breeding. We analyzed a diversity panel largely consisted of advanced lines derived from synthetic hexaploid wheats for allelic variation at 87 functional genes or loci of breeding importance using 124 high-throughput KASP markers. We also developed two KASP markers for water-soluble carbohydrate genes (<i>TaSST-D1</i> and <i>TaSST-A1</i>) associated with plant height and thousand grain weight (TGW) in the diversity panel. KASP genotyping results indicated that beneficial alleles for genes underpinning flowering time (<i>Ppd-D1</i> and <i>Vrn-D3</i>), thousand grain weight (<i>TaCKX-D1, TaTGW6-A1, TaSus1-7B</i>, and <i>TaCwi-D1</i>), water-soluble carbohydrates (<i>TaSST-A1</i>), yellow-pigment content (<i>Psy-B1</i> and <i>Zds-D1</i>), and root lesion nematodes (<i>Rlnn1</i>) were fixed in diversity panel with frequency ranged from 96.4 to 100%. The association analysis of functional genes with agronomic and biochemical traits under well-watered (WW) and water-limited (WL) conditions revealed that 21 marker-trait associations (MTAs) were consistently detected in both moisture conditions. The major developmental genes such as <i>Vrn-A1, Rht-D1</i>, and <i>Ppd-B1</i> had the confounding effect on several agronomic traits including plant height, grain size and weight, and grain yield in both WW and WL conditions. The accumulation of favorable alleles for grain size and weight genes additively enhanced grain weight in the diversity panel. Graphical genotyping approach was used to identify accessions with maximum number of favorable alleles, thus likely to have high breeding value. These results improved our knowledge on the selection of favorable and unfavorable alleles through unconscious selection breeding and identified the opportunities to deploy alleles with effects in wheat breeding.
Project description:MiR399 and its target PHOSPHATE2 (PHO2) play pivotal roles in phosphate signaling in plants. Loss of function mutation in PHO2 leads to excessive Pi accumulation in shoots and growth retardation in diploid plants like Arabidopsis thaliana and rice (Oryza sativa). Here we isolated three PHO2 homologous genes TaPHO2-A1, -B1 and -D1 from hexaploid wheat (Triticum aestivum). These TaPHO2 genes all contained miR399-binding sites and were able to be degraded by tae-miR399. TaPHO2-D1 was expressed much more abundantly than TaPHO2-A1 and -B1. The ion beam-induced deletion mutants were used to analyze the effects of TaPHO2s on phosphorus uptake and plant growth. The tapho2-a1, tapho2-b1 and tapho2-d1 mutants all had significant higher leaf Pi concentrations than did the wild type, with tapho2-d1 having the strongest effect, and tapho2-b1 the weakest. Two consecutive field experiments showed that knocking out TaPHO2-D1 reduced plant height and grain yield under both low and high phosphorus conditions. However, knocking out TaPHO2-A1 significantly increased phosphorus uptake and grain yield under low phosphorus conditions, with no adverse effect on grain yield under high phosphorus conditions. Our results indicated that TaPHO2s involved in phosphorus uptake and translocation, and molecular engineering TaPHO2 shows potential in improving wheat yield with less phosphorus fertilizer.
Project description:TaGS5 genes were cloned from bread wheat and were physically mapped on 3AS and 3DS. Sequencing results revealed that a SNP was found in the sixth exon of TaGS5-A1 gene. The SNP resulted in amino acid change from alanine to serine at the 303 bp position of TaGS5-A1. These two alleles were designated as TaGS5-A1a (alanine at the 303 bp position) and TaGS5-A1b genes (serine at the 303-bp position). Analysis of association of TaGS5-A1 alleles with agronomic traits indicated that cultivars with TaGS5-A1b possessed wider kernel width and higher thousand-kernel weight, as well as significantly lower plant height, spike length, and internode length below spike than those of cultivars with TaGS5-A1a over 3 years. These trait differences between TaGS5-A1a and TaGS5-A1b genotypes were larger in landraces than in modern cultivars. This finding suggested that TaGS5 gene played an important role in modulating yield-related traits in the landraces, which possibly resulted from numerous superior genes gathering in modern cultivars after strong artificial selection. The preferred TaGS5-A1b haplotype underwent very strong positive selection in Chinese modern wheat breeding, but not in Chinese landraces. Expression analysis of the TaGS5-A1 gene indicated that TaGS5-A1b allele possessed significantly higher expression level than TaGS5-A1b allele in differently developmental seeds. This study could provide relatively superior genotype in view of agronomic traits in wheat breeding programs. Likewise, this study could offer important information for the dissection of molecular and genetic basis of yield-related traits.