Genome-wide association study identifies novel loci and candidate genes for rust resistance in wheat (Triticum aestivum L.).

BACKGROUND: Wheat rusts are important biotic stresses, development of rust resistant cultivars through molecular approaches is both economical and sustainable. Extensive phenotyping of large mapping populations under diverse production conditions and high-density genotyping would be the ideal strategy to identify major genomic regions for rust resistance in wheat. The genome-wide association study (GWAS) population of 280 genotypes was genotyped using a 35 K Axiom single nucleotide polymorphism (SNP) array and phenotyped at eight, 10, and, 10 environments, respectively for stem/black rust (SR), stripe/yellow rust (YR), and leaf/brown rust (LR). RESULTS: Forty-one Bonferroni corrected marker-trait associations (MTAs) were identified, including 17 for SR and 24 for YR. Ten stable MTAs and their best combinations were also identified. For YR, AX-94990952 on 1A + AX-95203560 on 4A + AX-94723806 on 3D + AX-95172478 on 1A showed the best combination with an average co-efficient of infection (ACI) score of 1.36. Similarly, for SR, AX-94883961 on 7B + AX-94843704 on 1B and AX-94883961 on 7B + AX-94580041 on 3D + AX-94843704 on 1B showed the best combination with an ACI score of around 9.0. The genotype PBW827 have the best MTA combinations for both YR and SR resistance. In silico study identifies key prospective candidate genes that are located within MTA regions. Further, the expression analysis revealed that 18 transcripts were upregulated to the tune of more than 1.5 folds including 19.36 folds (TraesCS3D02G519600) and 7.23 folds (TraesCS2D02G038900) under stress conditions compared to the control conditions. Furthermore, highly expressed genes in silico under stress conditions were analyzed to find out the potential links to the rust phenotype, and all four genes were found to be associated with the rust phenotype. CONCLUSION: The identified novel MTAs, particularly stable and highly expressed MTAs are valuable for further validation and subsequent application in wheat rust resistance breeding. The genotypes with favorable MTA combinations can be used as prospective donors to develop elite cultivars with YR and SR resistance.

Integrated genomic selection for rapid improvement of crops.

An increase in the rate of crop improvement is essential for achieving sustained food production and other needs of ever-increasing population. Genomic selection (GS) is a potential breeding tool that has been successfully employed in animal breeding and is being incorporated into plant breeding. GS promises accelerated breeding cycles through a rapid selection of superior genotypes. Numerous empirical and simulation studies on GS and realized impacts on improvement in the crop yields are recently being reported. For a holistic understanding of the technology, we briefly discuss the concept of genetic gain, GS methodology, its current status, advantages of GS over other breeding methods, prediction models, and the factors controlling prediction accuracy in GS. Also, integration of speed breeding and other novel technologies viz. high throughput genotyping and phenotyping technologies for enhancing the efficiency and pace of GS, followed by its prospective applications in varietal development programs is reviewed.

Optimization of Agrobacterium-mediated transformation in spring bread wheat using mature and immature embryos.

Wheat is the most widely grown staple food crop in the world and accounts for dietary needs of more than 35% of the human population. Current status of transgenic wheat development is slow all over the world due to the lack of a suitable transformation system. In the present study, an efficient and reproducible Agrobacterium-mediated transformation system in bread wheat (Triticum aestivum L.) is established. The mature and immature embryos of six recently released high yielding spring bread wheat genotypes were used to standardize various parameters using Agrobacterium tumefaciens strain EHA105 harbouring binary vector pCAMBIA3301 having gus and bar as marker genes. The optimum duration for embryo pre-culture, inoculation time and co-cultivation were 2 days, 30 min and 48 h, respectively. The bacterial inoculum concentration of OD of 1 at 600 nm showed 67.25% transient GUS expression in the histochemical GUS assay. The filter paper based co-cultivation limits the Agrobacterium overgrowth and had 82.3% explants survival rate whereas medium based strategy had 22.7% explants survival only. The medium having picloram 4 mg/l along with antibiotics (cefotaxime 500 mg/l and timentin 300 mg/l) was found best suitable for initial week callus induction. The standardized procedure gave overall 14.9% transformation efficiency in immature embryos and 9.8% in mature embryos and confirmed by gene-specific and promoter-specific PCR and southern analysis. These results indicate that the developed Agrobacterium-mediated transformation system is suitable for diverse wheat genotypes. The major obstacle for the implication of the CRISPR-based genome editing techniques is the non-availability of a suitable transformation system. Thus, the present system can be exploited to deliver the T-DNA into the wheat genome for CRISPR-based target modifications and transgene insertions.

Development of an efficient and reproducible regeneration system in wheat (Triticum aestivum L.).

The availability of reproducible regeneration system through tissue culture is a major bottleneck in wheat improvement program. The present study has considered to develop an efficient callus induction and regeneration system using mature and immature embryos as explants in recently released agronomically superior spring wheat varieties. An efficient sterilization process was standardized using 0.1% HgCl2 and 70% ethanol for both seeds and embryos. The maximum possible combinations of plant growth regulators (PGRs) were evaluated for their effect on different wheat regeneration processes through tissue culture starting from callus to root induction. Picloram is found as an effective auxin with 87.63-98.67% callus induction efficiency in both explants. Supplementation of CuSO4 along with 2,4-D, zeatin in regeneration medium significantly enhanced the multiple shoot induction. The shoot development was achieved using full strength Murashige and Skoog's (MS) medium and root induction using half MS medium without PGRs. The optimized medium and method has resulted up to 100% regeneration irrespective of the genotype used with high reproducibility. Thus, the standardized regeneration system can be used in the regeneration of healthy plants from embryos rescued from interspecies crosses, transgenic production, induced mutation breeding and recently developed genome editing techniques for the procreation of wheat plants having novel traits.

Genome-Wide Association Study for Grain Protein, Thousand Kernel Weight, and Normalized Difference Vegetation Index in Bread Wheat (Triticum aestivum L.).

Genomic regions governing grain protein content (GPC), 1000 kernel weight (TKW), and normalized difference vegetation index (NDVI) were studied in a set of 280 bread wheat genotypes. The genome-wide association (GWAS) panel was genotyped using a 35K Axiom array and phenotyped in three environments. A total of 26 marker-trait associations (MTAs) were detected on 18 chromosomes covering the A, B, and D subgenomes of bread wheat. The GPC showed the maximum MTAs (16), followed by NDVI (6), and TKW (4). A maximum of 10 MTAs was located on the B subgenome, whereas, 8 MTAs each were mapped on the A and D subgenomes. In silico analysis suggest that the SNPs were located on important putative candidate genes such as NAC domain superfamily, zinc finger RING-H2-type, aspartic peptidase domain, folylpolyglutamate synthase, serine/threonine-protein kinase LRK10, pentatricopeptide repeat, protein kinase-like domain superfamily, cytochrome P450, and expansin. These candidate genes were found to have different roles including regulation of stress tolerance, nutrient remobilization, protein accumulation, nitrogen utilization, photosynthesis, grain filling, mitochondrial function, and kernel development. The effects of newly identified MTAs will be validated in different genetic backgrounds for further utilization in marker-aided breeding.

Harnessing High Yield Potential in Wheat (Triticum aestivum L.) under Climate Change Scenario.

Wheat is a major staple food crop for food security in India and South Asia. The current rate (0.8-1.2%) of genetic gain in wheat is significantly shorter than the 2.4% needed to meet future demand. The changing climate and increased yield loss due to factors such as terminal heat stress necessitate the need for climate-resilient practices to sustain wheat production. At ICAR-Indian Institute of Wheat and Barley Research in Karnal, Haryana, India, a new High Yield Potential Trial (HYPT) was conceptualized and subsequently conducted at six locations in the highly productive North Western Plain Zone (NWPZ). An attempt was made to harness higher wheat yields through the best pipeline genotypes suitable for early sowing and modified agronomic practices to explore the feasibility of a new approach that is profitable to farmers. The modified agronomic practices included like early sowing, application of 150% recommended dose of fertilizers, and two sprays of growth regulators (Chlormaquate chloride and Tebuconazole) to prevent lodging. The mean yield in the HYPT was 19.4% superior compared to the best trials conducted during the normal sowing time. A highly positive and significant correlation of grain yield with grain filling duration (0.51), biomass (0.73), harvest index (0.75), normalized difference vegetation Index (0.27), chlorophyll content index (0.32), and 1000-grain weight (0.62) was observed. An increased return of USD 201.95/ha was realized in the HYPT when compared to normal sowing conditions. This study proves that new integrated practices have the potential to provide the best profitable yields in wheat in the context of climate change.

Genetic dissection of grain iron and zinc, and thousand kernel weight in wheat (Triticum aestivum L.) using genome-wide association study.

Genetic biofortification is recognized as a cost-effective and sustainable strategy to reduce micronutrient malnutrition. Genomic regions governing grain iron concentration (GFeC), grain zinc concentration (GZnC), and thousand kernel weight (TKW) were investigated in a set of 280 diverse bread wheat genotypes. The genome-wide association (GWAS) panel was genotyped using 35 K Axiom Array and phenotyped in five environments. The GWAS analysis showed a total of 17 Bonferroni-corrected marker-trait associations (MTAs) in nine chromosomes representing all the three wheat subgenomes. The TKW showed the highest MTAs (7), followed by GZnC (5) and GFeC (5). Furthermore, 14 MTAs were identified with more than 10% phenotypic variation. One stable MTA i.e. AX-95025823 was identified for TKW in both E4 and E5 environments along with pooled data, which is located at 68.9 Mb on 6A chromosome. In silico analysis revealed that the SNPs were located on important putative candidate genes such as Multi antimicrobial extrusion protein, F-box domain, Late embryogenesis abundant protein, LEA-18, Leucine-rich repeat domain superfamily, and C3H4 type zinc finger protein, involved in iron translocation, iron and zinc homeostasis, and grain size modifications. The identified novel MTAs will be validated to estimate their effects in different genetic backgrounds for subsequent use in marker-assisted selection. The identified SNPs will be valuable in the rapid development of biofortified wheat varieties to ameliorate the malnutrition problems.