Corrigendum: Unravelling the genetic framework associated with grain quality and yield-related traits in maize (Zea mays L.).

[This corrects the article DOI: 10.3389/fgene.2023.1248697.].

Integrating Association Mapping, Linkage Mapping, Fine Mapping with RNA Seq Conferring Seedling Vigor Improvement for Successful Crop Establishment in Deep Sown Direct-Seeded Rice.

BACKGROUND: Ongoing large-scale shift towards direct seeded rice (DSR) necessitates a convergence of breeding and genetic approaches for its sustenance and harnessing natural resources and environmental benefits. Improving seedling vigour remains key objective for breeders working with DSR. The present study aims to understand the genetic control of seedling vigour in deep sown DSR. Combined genome-wide association mapping, linkage mapping, fine mapping, RNA-sequencing to identify candidate genes and validation of putative candidate genes were performed in the present study. RESULTS: Significant phenotypic variations were observed among genotypes in both F3:4:5 and BC2F2:3 populations. The mesocotyl length showed significant positive correlation with %germination, root and shoot length. The 881 kb region on chromosome 7 reported to be associated with mesocotyl elongation. RNA-seq data and RT-PCR results identified and validated seven potential candidate genes. The four promising introgression lines free from linkage drag and with longer mesocotyl length, longer root length, semi-dwarf plant height have been identified. CONCLUSION: The study will provide rice breeders (1) the pre breeding material in the form of anticipated DSR adapted introgression lines possessing useful traits and alleles improving germination under deep sown DSR field conditions (2) the base for the studies involving functional characterization of candidate genes. The development and utilization of improved introgression lines and molecular markers may play an important role in genomics-assisted breeding (GAB) during the pyramiding of valuable genes providing adaptation to rice under DSR. Our results offer a robust and reliable package that can contribute towards enhancing genetic gains in direct seeded rice breeding programs.

Unravelling the genetic framework associated with grain quality and yield-related traits in maize (Zea mays L.).

Maize serves as a crucial nutrient reservoir for a significant portion of the global population. However, to effectively address the growing world population's hidden hunger, it is essential to focus on two key aspects: biofortification of maize and improving its yield potential through advanced breeding techniques. Moreover, the coordination of multiple targets within a single breeding program poses a complex challenge. This study compiled mapping studies conducted over the past decade, identifying quantitative trait loci associated with grain quality and yield related traits in maize. Meta-QTL analysis of 2,974 QTLs for 169 component traits (associated with quality and yield related traits) revealed 68 MQTLs across different genetic backgrounds and environments. Most of these MQTLs were further validated using the data from genome-wide association studies (GWAS). Further, ten MQTLs, referred to as breeding-friendly MQTLs (BF-MQTLs), with a significant phenotypic variation explained over 10% and confidence interval less than 2 Mb, were shortlisted. BF-MQTLs were further used to identify potential candidate genes, including 59 genes encoding important proteins/products involved in essential metabolic pathways. Five BF-MQTLs associated with both quality and yield traits were also recommended to be utilized in future breeding programs. Synteny analysis with wheat and rice genomes revealed conserved regions across the genomes, indicating these hotspot regions as validated targets for developing biofortified, high-yielding maize varieties in future breeding programs. After validation, the identified candidate genes can also be utilized to effectively model the plant architecture and enhance desirable quality traits through various approaches such as marker-assisted breeding, genetic engineering, and genome editing.

Self-incompatibility: a targeted, unexplored pre-fertilization barrier in flower crops of Asteraceae.

Asteraceae (synonym as Compositae) is one of the largest angiosperm families among flowering plants comprising one-tenth of all agri-horticultural species grown across various habitats except in Antarctica. These are commercially utilized as cut and loose flowers as well as pot and bedding plants in landscape gardens due to their unique floral traits. Consequently, ineffective seed setting and presence of an intraspecific reproductive barrier known as self-incompatibility (SI) severely reduces the effectiveness of hybridization and self-fertilization by traditional crossing. There have been very few detailed studies of pollen-stigma interactions in this family. Moreover, about 63% of Aster species can barely self-fertilize due to self-incompatibility (SI). The chrysanthemum (Chrysanthemum × morifolium) is one of the most economically important ornamental plants in the Asteraceae family which hugely shows incompatibility. Reasons for the low fertility and reproductive capacity of species are still indefinite or not clear. Hence, the temporal pattern of inheritance of self-incompatibility and its effect on reproductive biology needs to be investigated further to improve the breeding efficiency. This review highlights the self-incompatible (SI) system operating in important Astraceous (ornamental) crops which are adversely affected by this mechanism along with different physiological and molecular techniques involved in breaking down self-incompatibility.

Development and validation of a novel core set of KASP markers for the traits improving grain yield and adaptability of rice under direct-seeded cultivation conditions.

The development and utilization of molecular-markers play an important role in genomics-assisted breeding during pyramiding of valuable genes. The aim of present study was to develop and validate a novel core-set of KASP (Kompetitive Allele-Specific PCR) markers associated with traits improving rice grain yield and adaptability under direct-seeded cultivation conditions. The 110 phenotypically validated KASP assays out of 171 designed KASP, include assays for biotic-resistance genes, anaerobic germination, root-traits, grain yield, lodging resistance and early-uniform emergence. The KASP assays were validated for their robustness and reliability at five different levels using diverse germplasm, segregating and advanced population, comparison with SSR markers and on F1s. The present research work will provide (i) breeding material in form of anticipated pre-direct-seeded adapted rice varieties (ii) single improved breeding line with many useful genes and (iii) KASP assay information for the useful QTL/genes providing grain yield and adaptability to rice under direct-seeded cultivation conditions.

Genome wide association analyses to understand genetic basis of flowering and plant height under three levels of nitrogen application in Brassica juncea (L.) Czern & Coss.

Timely transition to flowering, maturity and plant height are important for agronomic adaptation and productivity of Indian mustard (B. juncea), which is a major edible oilseed crop of low input ecologies in Indian subcontinent. Breeding manipulation for these traits is difficult because of the involvement of multiple interacting genetic and environmental factors. Here, we report a genetic analysis of these traits using a population comprising 92 diverse genotypes of mustard. These genotypes were evaluated under deficient (N75), normal (N100) or excess (N125) conditions of nitrogen (N) application. Lower N availability induced early flowering and maturity in most genotypes, while high N conditions delayed both. A genotyping-by-sequencing approach helped to identify 406,888 SNP markers and undertake genome wide association studies (GWAS). 282 significant marker-trait associations (MTA's) were identified. We detected strong interactions between GWAS loci and nitrogen levels. Though some trait associated SNPs were detected repeatedly across fertility gradients, majority were identified under deficient or normal levels of N applications. Annotation of the genomic region (s) within ± 50 kb of the peak SNPs facilitated prediction of 30 candidate genes belonging to light perception, circadian, floral meristem identity, flowering regulation, gibberellic acid pathways and plant development. These included over one copy each of AGL24, AP1, FVE, FRI, GID1A and GNC. FLC and CO were predicted on chromosomes A02 and B08 respectively. CDF1, CO, FLC, AGL24, GNC and FAF2 appeared to influence the variation for plant height. Our findings may help in improving phenotypic plasticity of mustard across fertility gradients through marker-assisted breeding strategies.

Meta-QTL Analysis in Rice and Cross-Genome Talk of the Genomic Regions Controlling Nitrogen Use Efficiency in Cereal Crops Revealing Phylogenetic Relationship.

The phenomenal increase in the use of nitrogenous fertilizers coupled with poor nitrogen use efficiency is among the most important threats to the environment, economic, and social health. During the last 2 decades, a number of genomic regions associated with nitrogen use efficiency (NUE) and related traits have been reported by different research groups, but none of the stable and major effect QTL have been utilized in the marker-assisted introgression/pyramiding program. Compiling the data available in the literature could be very useful in identifying stable and major effect genomic regions associated with the root and NUE-related trait improving the rice grain yield. In the present study, we performed meta-QTL analysis on 1,330 QTL from 29 studies published in the past 2 decades. A total of 76 MQTL with a stable effect over different genetic backgrounds and environments were identified. The significant reduction in the confidence interval of the MQTL compared to the initial QTL resulted in the identification of annotated and putative candidate genes related to the traits considered in the present study. A hot spot region associated with correlated traits on chr 1, 4, and 8 and candidate genes associated with nitrate transporters, nitrogen content, and ammonium uptake on chromosomes 2, 4, 6, and 8 have been identified. The identified MQTL, putative candidate genes, and their orthologues were validated on our previous studies conducted on rice and wheat. The research-based interventions such as improving nitrogen use efficiency via identification of major genomic regions and candidate genes can be a plausible, simple, and low-cost solution to address the challenges of the crop improvement program.

Association genetics of the parameters related to nitrogen use efficiency in Brassica juncea L.

KEY MESSAGE: Genome wide association studies allowed prediction of 17 candidate genes for association with nitrogen use efficiency. Novel information obtained may provide better understanding of genomic controls underlying germplasm variations for this trait in Indian mustard. Nitrogen use efficiency (NUE) of Indian mustard (Brassica juncea (L.) Czern & Coss.) is low and most breeding efforts to combine NUE with crop performance have not succeeded. Underlying genetics also remain unexplored. We tested 92 SNP-genotyped inbred lines for yield component traits, N uptake efficiency (NUPEFF), nitrogen utilization efficiency (NUTEFF), nitrogen harvest index (NHI) and NUE for two years at two nitrogen doses (No without added N and N100 added @100 kg/ha). Genotypes IC-2489-88, M-633, MCP-632, HUJM 1080, GR-325 and DJ-65 recorded high NUE at low N. These also showed improved crop performance under high N. One determinate mustard genotype DJ-113 DT-3 revealed maximum NUTEFF. Genome wide association studies (GWAS) facilitated recognition of 17 quantitative trait loci (QTLs). Environment specificity was high. B-genome chromosomes (B02, B03, B05, B07 and B08) harbored many useful loci. We also used regional association mapping (RAM) to supplement results from GWAS. Annotation of the genomic regions around peak SNPs helped to predict several gene candidates for root architecture, N uptake, assimilation and remobilization. CAT9 (At1g05940) was consistently envisaged for both NUE and NUPEFF. Major N transporter genes, NRT1.8 and NRT3.1 were predicted for explaining variation for NUTEFF and NUPEFF, respectively. Most significant amino acid transporter gene, AAP1 appeared associated with NUE under limited N conditions. All these candidates were predicted in the regions of high linkage disequilibrium. Sequence information of the predicted candidate genes will permit development of molecular markers to aid breeding for high NUE.

Association Mapping of Seed Quality Traits Under Varying Conditions of Nitrogen Application in Brassica juncea L. Czern & Coss.

Indian mustard (Brassica juncea) is a major source of vegetable oil in the Indian subcontinent. The seed cake left after the oil extraction is used as livestock feed. We examined the genetic architecture of oil, protein, and glucosinolates by conducting a genome-wide association study (GWAS), using an association panel comprising 92 diverse genotypes. We conducted trait phenotyping over 2 years at two levels of nitrogen (N) application. Genotyping by sequencing was used to identify 66,835 loci, covering 18 chromosomes. Genetic diversity and phenotypic variations were high for the studied traits. Trait performances were stable when averaged over years and N levels. However, individual performances differed. General and mixed linear models were used to estimate the association between the SNP markers and the seed quality traits. Population structure, principal components (PCs) analysis, and discriminant analysis of principal components (DAPCs) were included as covariates to overcome the bias due to the population stratification. We identified 16, 23, and 27 loci associated with oil, protein, and glucosinolates, respectively. We also established LD patterns and haplotype structures for the candidate genes. The average block sizes were larger on A-genome chromosomes as compared to the B- genome chromosomes. Genetic associations differed over N levels. However, meta-analysis of GWAS datasets not only improved the power to recognize associations but also helped to identify common SNPs for oil and protein contents. Annotation of the genomic region around the identified SNPs led to the prediction of 21 orthologs of the functional candidate genes related to the biosynthesis of oil, protein, and glucosinolates. Notable among these are: LACS5 (A09), FAD6 (B05), ASN1 (A06), GTR2 (A06), CYP81G1 (B06), and MYB44 (B06). The identified loci will be very useful for marker-aided breeding for seed quality modifications in B. juncea.

Molecular and genetic analysis of defensive responses of Brassica juncea - B. fruticulosa introgression lines to Sclerotinia infection.

Sclerotinia stem rot caused by Sclerotinia sclerotiorum is a major disease of crop brassicas, with inadequate variation for resistance in primary gene pools. We utilized a wild Brassicaceae species with excellent resistance against stem rot to develop a set of B. juncea - B. fruticulosa introgression lines (ILs). These were assessed for resistance using a highly reproducible stem inoculation technique against a virulent pathogen isolate. Over 40% of ILs showed higher levels of resistance. IL-43, IL-175, IL-215, IL-223 and IL-277 were most resistant ILs over three crop seasons. Sequence reads (21x) from the three most diverse ILs were then used to create B. juncea pseudomolecules, by replacing SNPs of reference B. juncea with those of re-sequenced ILs. Genotyping by sequencing (GBS) was also carried out for 88 ILs. Resultant sequence tags were then mapped on to the B. juncea pseudomolecules, and SNP genotypes prepared for each IL. Genome wide association studies helped to map resistance responses to stem rot. A total of 13 significant loci were identified on seven B. juncea chromosomes (A01, A03, A04, A05, A08, A09 and B05). Annotation of the genomic region around identified SNPs allowed identification of 20 candidate genes belonging to major disease resistance protein families, including TIR-NBS-LRR class, Chitinase, Malectin/receptor-like protein kinase, defensin-like (DEFL), desulfoglucosinolate sulfotransferase protein and lipoxygenase. A majority of the significant SNPs could be validated using whole genome sequences (21x) from five advanced generation lines being bred for Sclerotinia resistance as compared to three susceptible B. juncea germplasm lines. Our findings not only provide critical new understanding of the defensive pathway of B. fruticulosa resistance, but will also enable development of marker candidates for assisted transfer of introgressed resistant loci in to agronomically superior cultivars of crop Brassica.

Detection of First Marker Trait Associations for Resistance Against Sclerotinia sclerotiorum in Brassica juncea-Erucastrum cardaminoides Introgression Lines.

A set of 96 Brassica juncea-Erucastrum cardaminoides introgression lines (ILs) were developed with genomic regions associated with Sclerotinia stem rot (Sclerotinia sclerotiorum) resistance from a wild Brassicaceous species E. cardaminoides. ILs were assessed for their resistance responses to stem inoculation with S. sclerotiorum, over three crop seasons (season I, 2011/2012; II, 2014/2015; III, 2016-2017). Initially, ILs were genotyped with transferable SSR markers and subsequently through genotyping by sequencing. SSR based association mapping identified six marker loci associated to resistance in both A and B genomes. Subsequent genome-wide association analysis (GWAS) of 84 ILs recognized a large number of SNPs associated to resistance, in chromosomes A03, A06, and B03. Chromosomes A03 and A06 harbored the maximum number of resistance related SNPs. Annotation of linked genomic regions highlighted an array of resistance mechanisms in terms of signal transduction pathways, hypersensitive responses and production of anti-fungal proteins and metabolites. Of major importance was the clustering of SNPs, encoding multiple resistance genes on small regions spanning approximately 885 kb region on chromosome A03 and 74 kb on B03. Five SNPs on chromosome A03 (6,390,210-381) were associated with LRR-RLK (receptor like kinases) genes that encode LRR-protein kinase family proteins. Genetic factors associated with pathogen-associated molecular patterns (PAMPs) and effector-triggered immunity (ETI) were predicted on chromosome A03, exhibiting 11 SNPs (6,274,763-994). These belonged to three R-Genes encoding TIR-NBS-LRR proteins. Marker trait associations (MTAs) identified will facilitate marker assisted introgression of these critical resistances, into new cultivars of B. juncea initially and, subsequently, into other crop Brassica species.