Identification of marker trait association is a prerequisite for marker-assisted breeding. To find markers linked with traits under heat and drought stress in bread wheat (Triticum aestivum L.), we performed a genome-wide association study (GWAS). GWAS mapping panel used in this study consists of advanced breeding lines from the IARI stress breeding programme produced by pairwise and complex crosses. Phenotyping was done at multi locations namely New Delhi, Karnal, Indore, Jharkhand and Pune with augmented-RCBD design under different moisture and heat stress regimes, namely timely sown irrigated (IR), timely sown restricted irrigated (RI) and late sown (LS) conditions. Yield and its component traits, viz., Days to Heading (DH), Days to Maturity (DM), Normalized Difference Vegetation Index (NDVI), Chlorophyll Content (SPAD), Canopy temperature (CT), Plant Height (PH), Thousand grain weight (TGW), Grain weight per spike (GWPS), Plot Yield (PLTY) and Biomass (BMS) were phenotyped. Analysis of variance and descriptive statistics revealed significant differences among the studied traits. Genotyping was done using the 35k SNP Wheat Breeder's Genotyping Array. Population structure and diversity analysis using filtered 10,546 markers revealed two subpopulations with sufficient diversity. A large whole genome LD block size of 7.15 MB was obtained at half LD decay value. Genome-wide association search identified 57 unique markers associated with various traits across the locations. Twenty-three markers were identified to be stable, among them nine pleiotropic markers were also identified. In silico search of the identified markers against the IWGSC ref genome revealed the presence of a majority of the SNPs at or near the gene coding region. These SNPs can be used for marker-assisted transfer of genes/QTLs after validation to develop climate-resilient cultivars.

Biofortification is gaining importance globally to improve human nutrition through enhancing the micronutrient content, such as vitamin A, iron, and zinc, in staple food crops. The present study aims to identify the chromosomal regions governing the grain iron concentration (GFeC), grain zinc concentration (GZnC), and thousand kernel weight (TKW) using recombinant inbred lines (RILs) in wheat, developed from a cross between HD3086 and HI1500. The experiment was conducted in four different production conditions at Delhi viz., control, drought, heat, and combined heat and drought stress and at Indore under drought stress. Grain iron and zinc content increased under heat and combined stress conditions, while thousand kernel weight decreased. Medium to high heritability with a moderate correlation between grain iron and zinc was observed. Out of 4,106 polymorphic markers between the parents, 3,407 SNP markers were used for linkage map construction which spanned over a length of 14791.18 cm. QTL analysis identified a total of 32 chromosomal regions governing the traits under study, which includes 9, 11, and 12 QTLs for GFeC, GZnC, and TKW, respectively. A QTL hotspot was identified on chromosome 4B which is associated with grain iron, grain zinc, and thousand kernel weight explaining the phenotypic variance of 29.28, 10.98, and 17.53%, respectively. Similarly, common loci were identified on chromosomes 4B and 4D for grain iron, zinc, and thousand kernel weight. In silico analysis of these chromosomal regions identified putative candidate genes that code for proteins such as Inositol 1,3,4-trisphosphate 5/6-kinase, P-loop containing nucleoside triphosphate hydrolase, Pleckstrin homology (PH) domains, Serine-threonine/tyrosine-protein kinase and F-box-like domain superfamily proteins which play role in many important biochemical or physiological process. The identified markers linked to QTLs can be used in MAS once successfully validated.

IntroductionWheat is grown and consumed worldwide, making it an important staple food crop for both its calorific and nutritional content. In places where wheat is used as a staple food, suboptimal micronutrient content levels, especially of grain iron (Fe) and zinc (Zn), can lead to malnutrition. Grain nutrient content is influenced by abiotic stresses, such as drought and heat stress. The best method for addressing micronutrient deficiencies is the biofortification of food crops. The prerequisites for marker-assisted varietal development are the identification of the genomic region responsible for high grain iron and zinc contents and an understanding of their genetics.MethodsA total of 193 diverse wheat genotypes were evaluated under drought and heat stress conditions across the years at the Indian Agricultural Research Institute (IARI), New Delhi, under timely sown irrigated (IR), restricted irrigated (RI) and late sown (LS) conditions. Grain iron content (GFeC) and grain zinc content (GZnC) were estimated from both the control and treatment groups. Genotyping of all the lines under study was carried out with the single nucleotide polymorphisms (SNPs) from Breeder’s 35K Axiom Array.Result and DiscussionThree subgroups were observed in the association panel based on both principal component analysis (PCA) and dendrogram analysis. A large whole-genome linkage disequilibrium (LD) block size of 3.49 Mb was observed. A genome-wide association study identified 16 unique stringent marker trait associations for GFeC, GZnC, and 1000-grain weight (TGW). In silico analysis demonstrated the presence of 28 potential candidate genes in the flanking region of 16 linked SNPs, such as synaptotagmin-like mitochondrial-lipid-binding domain, HAUS augmin-like complex, di-copper center-containing domain, protein kinase, chaperonin Cpn60, zinc finger, NUDIX hydrolase, etc. Expression levels of these genes in vegetative tissues and grain were also found. Utilization of identified markers in marker-assisted breeding may lead to the rapid development of biofortified wheat genotypes to combat malnutrition.

There is a significant yield reduction in the wheat crop as a result of different biotic and abiotic stresses, and changing climate, among them moisture deficit stress and leaf rust are the major ones affecting wheat worldwide. HD3086 is a high-yielding wheat variety that has been released for commercial cultivation under timely sown irrigated conditions in the Indo-Gangetic plains of India. Variety HD3086 provides a good, stable yield, and it is the choice of millions of farmers in India. It becomes susceptible to the most prevalent pathotypes 77-5 and 77-9 of Puccinia triticina (causing leaf rust) in the production environment and its potential yield cannot be realized under moisture deficit stress. The present study demonstrates the use of a marker-assisted back cross breeding approach to the successful transfer of leaf rust resistance gene Lr24 and QTLs linked to moisture deficit stress tolerance in the background of HD3086. The genotype HI1500 was used as a donor parent that possesses leaf rust-resistant gene Lr24, which confers resistance against the major pathotypes found in the production environment. It possesses inbuilt tolerance under abiotic stresses with superior quality traits. Foreground selection for gene Lr24 and moisture deficit stress tolerance QTLs linked to Canopy temperature (CT), Normal Differential Vegetation Index (NDVI) and Thousand Kernel Weight (TKW) in different generations of the backcrossing and selection. In BC2F2, foreground selection was carried out to identify homozygous lines based on the linked markers and were advanced following pedigree based phenotypic selection. The selected lines were evaluated against P. triticina pathotypes 77-5 and 77-9 under controlled conditions. Recurrent parent recovery of the selected lines ranged from 78-94%. The identified lines were evaluated for their tolerance to moisture stress under field conditions and their resistance to rust under artificial epiphytotic conditions for two years. In BC2F5 generation, eight positive lines for marker alleles were selected which showed resistance to leaf rust and recorded an improvement in component traits of moisture deficit stress tolerance such as CT, NDVI, TKW and yield compared to the recurrent parent HD3086. The derived line is named HD3471 and is nominated for national trials for testing and further release for commercial cultivation.

Wheat is the staple food crop of global importance for its grain nutrient quality. Grain iron and zinc content of the wheat grain is an important quantitatively inherited trait that is influenced by the environmental factors such as drought and heat stress. Phenotypic evaluation of 295 advanced breeding lines from the wheat stress breeding program of IARI was carried out under timely sown irrigated (IR), restricted irrigated, and late-sown conditions at New Delhi during the cropping season of 2020–21, and grain iron (GFeC) and zinc (GZnC) contents were estimated from both control and treatments. A statistically significant increase in GFeC and GZnC was observed under stress conditions compared to that of the control. Genotyping was carried out with the SNPs from the 35K Axiom Breeder’s array, and marker–trait association was identified by GWAS analysis. Of the 23 MTAs identified, seven were linked with GFeC and sixteen were linked with GZnC. In silico analysis revealed a few important transcripts involved in various plant metabolism, growth, and development activities such as auxin response factor, root UVB sensitive proteins, potassium transporter, glycosyl transferase, COBRA, and F-box-like domain. The identified MTAs can be used for molecular breeding after validation and also for rapid development of micronutrient-rich varieties of wheat to mitigate hidden hunger.