Aspergillus flavus pangenome (AflaPan) uncovers novel aflatoxin and secondary metabolite associated gene clusters.

BACKGROUND: Aspergillus flavus is an important agricultural and food safety threat due to its production of carcinogenic aflatoxins. It has high level of genetic diversity that is adapted to various environments. Recently, we reported two reference genomes of A. flavus isolates, AF13 (MAT1-2 and highly aflatoxigenic isolate) and NRRL3357 (MAT1-1 and moderate aflatoxin producer). Where, an insertion of 310 kb in AF13 included an aflatoxin producing gene bZIP transcription factor, named atfC. Observations of significant genomic variants between these isolates of contrasting phenotypes prompted an investigation into variation among other agricultural isolates of A. flavus with the goal of discovering novel genes potentially associated with aflatoxin production regulation. Present study was designed with three main objectives: (1) collection of large number of A. flavus isolates from diverse sources including maize plants and field soils; (2) whole genome sequencing of collected isolates and development of a pangenome; and (3) pangenome-wide association study (Pan-GWAS) to identify novel secondary metabolite cluster genes. RESULTS: Pangenome analysis of 346 A. flavus isolates identified a total of 17,855 unique orthologous gene clusters, with mere 41% (7,315) core genes and 59% (10,540) accessory genes indicating accumulation of high genomic diversity during domestication. 5,994 orthologous gene clusters in accessory genome not annotated in either the A. flavus AF13 or NRRL3357 reference genomes. Pan-genome wide association analysis of the genomic variations identified 391 significant associated pan-genes associated with aflatoxin production. Interestingly, most of the significantly associated pan-genes (94%; 369 associations) belonged to accessory genome indicating that genome expansion has resulted in the incorporation of new genes associated with aflatoxin and other secondary metabolites. CONCLUSION: In summary, this study provides complete pangenome framework for the species of Aspergillus flavus along with associated genes for pathogen survival and aflatoxin production. The large accessory genome indicated large genome diversity in the species A. flavus, however AflaPan is a closed pangenome represents optimum diversity of species A. flavus. Most importantly, the newly identified aflatoxin producing gene clusters will be a new source for seeking aflatoxin mitigation strategies and needs new attention in research.

Linkage-Mapping and Genome-wide Association Study Identified Two Peanut Late Leaf Spot Resistance Loci, PLLSR-1 and PLLSR-2, Using a Nested Association Mapping.

Identification of candidate genes and molecular markers for late leaf spot (LLS) disease resistance in peanut (Arachis hypogaea) has been a focus of molecular breeding for the U.S. industry funded peanut genome project. Efforts have been hindered by limited mapping resolution due to low levels of genetic recombination and marker density available in traditional biparental mapping populations. To address this, a multi-parental nested association mapping (NAM) population has been genotyped with the peanut 58 K SNP array and phenotyped for LLS severity in the field for three years. Joint linkage-based QTL mapping identified nine QTLs for LLS resistance with significant phenotypic variance explained (PVE) up to 47.7%. A genome-wide association study (GWAS) identified 13 SNPs consistently associated with LLS resistance. Two genomic regions harboring the consistent QTLs and SNPs were identified from 1,336 Kb to 1,520 Kb (184 Kb) on chromosome B02 and from 1,026.9 Kb to 1,793.2 Kb (767 Kb) on chromosome B03, designated as peanut late leaf spot resistance loci, PLLSR-1 and PLLSR-2, respectively. PLLSR-1 contains 10 NBS-LRR disease resistant genes. An NBS-LRR disease resistance gene Arahy.VKVT6A was also identified on homoeologous chromosome A02. PLLSR-2 contains five significant SNPs associated with five different genes encoding callose synthase, pollen defective in guidance protein, pentatricopeptide repeat (PPR), acyl-activating enzyme, and C2 GRAM domains-containing protein. This study highlights the power of multi-parent populations such as NAM for genetic mapping and marker-trait association studies in peanuts. Validation of these two LLS resistance loci will be needed for marker-assisted breeding.

Homoeologous recombination is recurrent in the nascent synthetic allotetraploid Arachis ipaënsis × Arachis correntina4x and its derivatives.

Genome instability in newly synthesized allotetraploids of peanut has breeding implications that have not been fully appreciated. Synthesis of wild species-derived neo-tetraploids offers the opportunity to broaden the gene pool of peanut; however, the dynamics among the newly merged genomes creates predictable and unpredictable variation. Selfed progenies from the neo-tetraploid Arachis ipaënsis × Arachis correntina (A. ipaënsis × A. correntina)4x and F1 hybrids and F2 progenies from crosses between A. hypogaea × [A. ipaënsis × A. correntina]4x were genotyped by the Axiom Arachis 48 K SNP array. Homoeologous recombination between the A. ipaënsis and A. correntina derived subgenomes was observed in the S0 generation. Among the S1 progenies, these recombined segments segregated and new events of homoeologous recombination emerged. The genomic regions undergoing homoeologous recombination segregated mostly disomically in the F2 progenies from A. hypogaea × [A. ipaënsis × A. correntina]4x crosses. New homoeologous recombination events also occurred in the F2 population, mostly found on chromosomes 03, 04, 05, and 06. From the breeding perspective, these phenomena offer both possibilities and perils; recombination between genomes increases genetic diversity, but genome instability could lead to instability of traits or even loss of viability within lineages.

Development and Genetic Characterization of Peanut Advanced Backcross Lines That Incorporate Root-Knot Nematode Resistance From Arachis stenosperma.

Crop wild species are increasingly important for crop improvement. Peanut (Arachis hypogaea L.) wild relatives comprise a diverse genetic pool that is being used to broaden its narrow genetic base. Peanut is an allotetraploid species extremely susceptible to peanut root-knot nematode (PRKN) Meloidogyne arenaria. Current resistant cultivars rely on a single introgression for PRKN resistance incorporated from the wild relative Arachis cardenasii, which could be overcome as a result of the emergence of virulent nematode populations. Therefore, new sources of resistance may be needed. Near-immunity has been found in the peanut wild relative Arachis stenosperma. The two loci controlling the resistance, present on chromosomes A02 and A09, have been validated in tetraploid lines and have been shown to reduce nematode reproduction by up to 98%. To incorporate these new resistance QTL into cultivated peanut, we used a marker-assisted backcrossing approach, using PRKN A. stenosperma-derived resistant lines as donor parents. Four cycles of backcrossing were completed, and SNP assays linked to the QTL were used for foreground selection. In each backcross generation seed weight, length, and width were measured, and based on a statistical analysis we observed that only one generation of backcrossing was required to recover the elite peanut's seed size. A populating of 271 BC3F1 lines was genome-wide genotyped to characterize the introgressions across the genome. Phenotypic information for leaf spot incidence and domestication traits (seed size, fertility, plant architecture, and flower color) were recorded. Correlations between the wild introgressions in different chromosomes and the phenotypic data allowed us to identify candidate regions controlling these domestication traits. Finally, PRKN resistance was validated in BC3F3 lines. We observed that the QTL in A02 and/or large introgression in A09 are needed for resistance. This present work represents an important step toward the development of new high-yielding and nematode-resistant peanut cultivars.

Publisher Correction: A recombination bin-map identified a major QTL for resistance to Tomato Spotted Wilt Virus in peanut (Arachis hypogaea).

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Nested-association mapping (NAM)-based genetic dissection uncovers candidate genes for seed and pod weights in peanut (Arachis hypogaea).

Multiparental genetic mapping populations such as nested-association mapping (NAM) have great potential for investigating quantitative traits and associated genomic regions leading to rapid discovery of candidate genes and markers. To demonstrate the utility and power of this approach, two NAM populations, NAM_Tifrunner and NAM_Florida-07, were used for dissecting genetic control of 100-pod weight (PW) and 100-seed weight (SW) in peanut. Two high-density SNP-based genetic maps were constructed with 3341 loci and 2668 loci for NAM_Tifrunner and NAM_Florida-07, respectively. The quantitative trait locus (QTL) analysis identified 12 and 8 major effect QTLs for PW and SW, respectively, in NAM_Tifrunner, and 13 and 11 major effect QTLs for PW and SW, respectively, in NAM_Florida-07. Most of the QTLs associated with PW and SW were mapped on the chromosomes A05, A06, B05 and B06. A genomewide association study (GWAS) analysis identified 19 and 28 highly significant SNP-trait associations (STAs) in NAM_Tifrunner and 11 and 17 STAs in NAM_Florida-07 for PW and SW, respectively. These significant STAs were co-localized, suggesting that PW and SW are co-regulated by several candidate genes identified on chromosomes A05, A06, B05, and B06. This study demonstrates the utility of NAM population for genetic dissection of complex traits and performing high-resolution trait mapping in peanut.

A recombination bin-map identified a major QTL for resistance to Tomato Spotted Wilt Virus in peanut (Arachis hypogaea).

Tomato spotted wilt virus (TSWV) is a devastating disease to peanut growers in the South-eastern region of the United States. Newly released peanut cultivars in recent years are crucial as they have some levels of resistance to TSWV. One mapping population of recombinant inbred line (RIL) used in this study was derived from peanut lines of SunOleic 97R and NC94022. A whole genome re-sequencing approach was used to sequence these two parents and 140 RILs. A recombination bin-based genetic map was constructed, with 5,816 bins and 20 linkage groups covering a total length of 2004 cM. Using this map, we identified three QTLs which were colocalized on chromosome A01. One QTL had the largest effect of 36.51% to the phenotypic variation and encompassed 89.5 Kb genomic region. This genome region had a cluster of genes, which code for chitinases, strictosidine synthase-like, and NBS-LRR proteins. SNPs linked to this QTL were used to develop Kompetitive allele specific PCR (KASP) markers, and the validated KASP markers showed expected segregation of alleles coming from resistant and susceptible parents within the population. Therefore, this bin-map and QTL associated with TSWV resistance made it possible for functional gene mapping, map-based cloning, and marker-assisted breeding. This study identified the highest number of SNP makers and demonstrated recombination bin-based map for QTL identification in peanut. The chitinase gene clusters and NBS-LRR disease resistance genes in this region suggest the possible involvement in peanut resistance to TSWV.

A new source of root-knot nematode resistance from Arachis stenosperma incorporated into allotetraploid peanut (Arachis hypogaea).

Root-knot nematode is a very destructive pathogen, to which most peanut cultivars are highly susceptible. Strong resistance is present in the wild diploid peanut relatives. Previously, QTLs controlling nematode resistance were identified on chromosomes A02, A04 and A09 of Arachis stenosperma. Here, to study the inheritance of these resistance alleles within the genetic background of tetraploid peanut, an F2 population was developed from a cross between peanut and an induced allotetraploid that incorporated A. stenosperma, [Arachis batizocoi x A. stenosperma]4×. This population was genotyped using a SNP array and phenotyped for nematode resistance. QTL analysis allowed us to verify the major-effect QTL on chromosome A02 and a secondary QTL on A09, each contributing to a percentage reduction in nematode multiplication up to 98.2%. These were validated in selected F2:3 lines. The genome location of the large-effect QTL on A02 is rich in genes encoding TIR-NBS-LRR protein domains that are involved in plant defenses. We conclude that the strong resistance to RKN, derived from the diploid A. stenosperma, is transferrable and expressed in tetraploid peanut. Currently it is being used in breeding programs for introgressing a new source of nematode resistance and to widen the genetic basis of agronomically adapted peanut lines.

Major QTLs for Resistance to Early and Late Leaf Spot Diseases Are Identified on Chromosomes 3 and 5 in Peanut (Arachis hypogaea).

Early and late leaf spots (LLSs) are the major foliar diseases of peanut responsible for severely decreased yield in the absence of intensive fungicide spray programs. Pyramiding host resistance to leaf spots in elite cultivars is a sustainable solution to mitigate the diseases. In order to determine the genetic control of leaf spot disease resistance in peanut, a recombinant inbred line population (Florida-07 × GP-NC WS16) segregating for resistance to both diseases was used to construct a SNP-based linkage map consisting of 855 loci. QTL mapping revealed three resistance QTLs for LLS qLLSA05 (phenotypic variation explained, PVE = 7-10%), qLLSB03 (PVE = 5-7%), and qLLSB05 (PVE = 15-41%) that were consistently expressed over multi-year analysis. Two QTL, qLLSA05 and qLLSB05, confirmed our previously published QTL-seq results. For early leaf spot, three resistance QTLs were identified in multiple years, two on chromosome A03 (PVE = 8-12%) and one on chromosome B03 (PVE = 13-20%), with the locus qELSA03_1.1 coinciding with the previously published genomic region for LLS resistance in GPBD4. Comparative analysis of the genomic regions spanning the QTLs suggests that resistance to early and LLSs are largely genetically independent. In addition, QTL analysis on yield showed that the presence of resistance allele in qLLSB03 and qLLSB05 loci might result in protection from yield loss caused by LLS disease damage. Finally, post hoc analysis of the RIL subpopulation that was not utilized in the QTL mapping revealed that the flanking markers for these QTLs can successfully select for resistant and susceptible lines, confirming the effectiveness of pyramiding these resistance loci to improve host-plant resistance in peanut breeding programs using marker-assisted selection.

The genome sequence of segmental allotetraploid peanut Arachis hypogaea.

Like many other crops, the cultivated peanut (Arachis hypogaea L.) is of hybrid origin and has a polyploid genome that contains essentially complete sets of chromosomes from two ancestral species. Here we report the genome sequence of peanut and show that after its polyploid origin, the genome has evolved through mobile-element activity, deletions and by the flow of genetic information between corresponding ancestral chromosomes (that is, homeologous recombination). Uniformity of patterns of homeologous recombination at the ends of chromosomes favors a single origin for cultivated peanut and its wild counterpart A. monticola. However, through much of the genome, homeologous recombination has created diversity. Using new polyploid hybrids made from the ancestral species, we show how this can generate phenotypic changes such as spontaneous changes in the color of the flowers. We suggest that diversity generated by these genetic mechanisms helped to favor the domestication of the polyploid A. hypogaea over other diploid Arachis species cultivated by humans.

High-density genetic map using whole-genome resequencing for fine mapping and candidate gene discovery for disease resistance in peanut.

Whole-genome resequencing (WGRS) of mapping populations has facilitated development of high-density genetic maps essential for fine mapping and candidate gene discovery for traits of interest in crop species. Leaf spots, including early leaf spot (ELS) and late leaf spot (LLS), and Tomato spotted wilt virus (TSWV) are devastating diseases in peanut causing significant yield loss. We generated WGRS data on a recombinant inbred line population, developed a SNP-based high-density genetic map, and conducted fine mapping, candidate gene discovery and marker validation for ELS, LLS and TSWV. The first sequence-based high-density map was constructed with 8869 SNPs assigned to 20 linkage groups, representing 20 chromosomes, for the 'T' population (Tifrunner × GT-C20) with a map length of 3120 cM and an average distance of 1.45 cM. The quantitative trait locus (QTL) analysis using high-density genetic map and multiple season phenotyping data identified 35 main-effect QTLs with phenotypic variation explained (PVE) from 6.32% to 47.63%. Among major-effect QTLs mapped, there were two QTLs for ELS on B05 with 47.42% PVE and B03 with 47.38% PVE, two QTLs for LLS on A05 with 47.63% and B03 with 34.03% PVE and one QTL for TSWV on B09 with 40.71% PVE. The epistasis and environment interaction analyses identified significant environmental effects on these traits. The identified QTL regions had disease resistance genes including R-genes and transcription factors. KASP markers were developed for major QTLs and validated in the population and are ready for further deployment in genomics-assisted breeding in peanut.

Insight into Genes Regulating Postharvest Aflatoxin Contamination of Tetraploid Peanut from Transcriptional Profiling.

Postharvest aflatoxin contamination is a challenging issue that affects peanut quality. Aflatoxin is produced by fungi belonging to the Aspergilli group, and is known as an acutely toxic, carcinogenic, and immune-suppressing class of mycotoxins. Evidence for several host genetic factors that may impact aflatoxin contamination has been reported, e.g., genes for lipoxygenase (PnLOX1 and PnLOX2/PnLOX3 that showed either positive or negative regulation with Aspergillus infection), reactive oxygen species, and WRKY (highly associated with or differentially expressed upon infection of maize with Aspergillus flavus); however, their roles remain unclear. Therefore, we conducted an RNA-sequencing experiment to differentiate gene response to the infection by A. flavus between resistant (ICG 1471) and susceptible (Florida-07) cultivated peanut genotypes. The gene expression profiling analysis was designed to reveal differentially expressed genes in response to the infection (infected vs. mock-treated seeds). In addition, the differential expression of the fungal genes was profiled. The study revealed the complexity of the interaction between the fungus and peanut seeds as the expression of a large number of genes was altered, including some in the process of plant defense to aflatoxin accumulation. Analysis of the experimental data with "keggseq," a novel designed tool for Kyoto Encyclopedia of Genes and Genomes enrichment analysis, showed the importance of α-linolenic acid metabolism, protein processing in the endoplasmic reticulum, spliceosome, and carbon fixation and metabolism pathways in conditioning resistance to aflatoxin accumulation. In addition, coexpression network analysis was carried out to reveal the correlation of gene expression among peanut and fungal genes. The results showed the importance of WRKY, toll/Interleukin1 receptor-nucleotide binding site leucine-rich repeat (TIR-NBS-LRR), ethylene, and heat shock proteins in the resistance mechanism.

Variability of arginine content and yield components in Valencia peanut germplasm.

Peanut seeds are rich in arginine, an amino acid that has several positive effects on human health. Establishing the genetic variability of arginine content in peanut will be useful for breeding programs that have high arginine as one of their goals. The objective of this study was to evaluate the variation of arginine content, pods/plant, seeds/pod, seed weight, and yield in Valencia peanut germplasm. One hundred and thirty peanut genotypes were grown under field condition for two years. A randomized complete block design with three replications was used for this study. Arginine content was analyzed in peanut seeds at harvest using spectrophotometry. Yield and yield components were recorded for each genotype. Significant differences in arginine content and yield components were found in the tested Valencia peanut germplasm. Arginine content ranged from 8.68-23.35 µg/g seed. Kremena was the best overall genotype of high arginine content, number of pods/plant, 100 seed weight and pod yield.

Gene expression profiling describes the genetic regulation of Meloidogyne arenaria resistance in Arachis hypogaea and reveals a candidate gene for resistance.

Resistance to root-knot nematode was introgressed into cultivated peanut Arachis hypogaea from a wild peanut relative, A. cardenasii and previously mapped to chromosome A09. The highly resistant recombinant inbred RIL 46 and moderately resistant RIL 48 were selected from a population with cv. Gregory (susceptible) and Tifguard (resistant) as female and male parents, respectively. RNA-seq analysis was performed on these four genotypes using root tissue harvested from root-knot nematode infected plants at 0, 3, 7 days after inoculation. Differential gene expression analysis provides evidence that root-knot nematodes modulate biological pathways involved in plant hormone, defense, cell signaling, cytoskeleton and cell wall metabolism in a susceptible reaction. Corresponding to resistance reaction, an effector-induced-immune response mediated by an R-gene was identified in Tifguard. Mapping of the introgressed region indicated that 92% of linkage group A09 was of A. cardenasii origin in Tifguard. RIL46 and RIL 48 possessed 3.6% and 83.5% of the introgression on A09, respectively. Within the small introgressed region carried by RIL 46, a constitutively expressed TIR-NBS-LRR gene was identified as the candidate for nematode resistance. Potential defense responsive pathways include effector endocytosis through clathrin-coated vesicle trafficking, defense signaling through membrane lipid metabolism and mucilage production.

Genetic Dissection of Novel QTLs for Resistance to Leaf Spots and Tomato Spotted Wilt Virus in Peanut (Arachis hypogaea L.).

Peanut is an important crop, economically and nutritiously, but high production cost is a serious challenge to peanut farmers as exemplified by chemical spray to control foliar diseases such as leaf spots and thrips, the vectors of tomato spotted wilt virus (TSWV). The objective of this research was to map the quantitative trait loci (QTLs) for resistance to leaf spots and TSWV in one recombinant inbred line (RIL) mapping population of "Tifrunner × GT-C20" for identification of linked markers for marker-assisted breeding. Here, we report the improved genetic linkage map with 418 marker loci with a marker density of 5.3 cM/loci and QTLs associated with multi-year (2010-2013) field phenotypes of foliar disease traits, including early leaf spot (ELS), late leaf spot (LLS), and TSWV. A total of 42 QTLs were identified with phenotypic variation explained (PVE) from 6.36 to 15.6%. There were nine QTLs for resistance to ELS, 22 QTLs for LLS, and 11 QTLs for TSWV, including six, five, and one major QTLs with PVE higher than 10% for resistance to each disease, respectively. Of the total 42 QTLs, 34 were mapped on the A sub-genome and eight mapped on the B sub-genome suggesting that the A sub-genome harbors more resistance genes than the B sub-genome. This genetic linkage map was also compared with two diploid peanut physical maps, and the overall co-linearity was 48.4% with an average co-linearity of 51.7% for the A sub-genome and 46.4% for the B sub-genome. The identified QTLs associated markers and potential candidate genes will be studied further for possible application in molecular breeding in peanut genetic improvement for disease resistance.

Genome-wide SNP Genotyping Resolves Signatures of Selection and Tetrasomic Recombination in Peanut.

Peanut (Arachis hypogaea; 2n = 4x = 40) is a nutritious food and a good source of vitamins, minerals, and healthy fats. Expansion of genetic and genomic resources for genetic enhancement of cultivated peanut has gained momentum from the sequenced genomes of the diploid ancestors of cultivated peanut. To facilitate high-throughput genotyping of Arachis species, 20 genotypes were re-sequenced and genome-wide single nucleotide polymorphisms (SNPs) were selected to develop a large-scale SNP genotyping array. For flexibility in genotyping applications, SNPs polymorphic between tetraploid and diploid species were included for use in cultivated and interspecific populations. A set of 384 accessions was used to test the array resulting in 54 564 markers that produced high-quality polymorphic clusters between diploid species, 47 116 polymorphic markers between cultivated and interspecific hybrids, and 15 897 polymorphic markers within A. hypogaea germplasm. An additional 1193 markers were identified that illuminated genomic regions exhibiting tetrasomic recombination. Furthermore, a set of elite cultivars that make up the pedigree of US runner germplasm were genotyped and used to identify genomic regions that have undergone positive selection. These observations provide key insights on the inclusion of new genetic diversity in cultivated peanut and will inform the development of high-resolution mapping populations. Due to its efficiency, scope, and flexibility, the newly developed SNP array will be very useful for further genetic and breeding applications in Arachis.

Mapping Quantitative Trait Loci of Resistance to Tomato Spotted Wilt Virus and Leaf Spots in a Recombinant Inbred Line Population of Peanut (Arachis hypogaea L.) from SunOleic 97R and NC94022.

Peanut is vulnerable to a range of diseases, such as Tomato spotted wilt virus (TSWV) and leaf spots which will cause significant yield loss. The most sustainable, economical and eco-friendly solution for managing peanut diseases is development of improved cultivars with high level of resistance. We developed a recombinant inbred line population from the cross between SunOleic 97R and NC94022, named as the S-population. An improved genetic linkage map was developed for the S-population with 248 marker loci and a marker density of 5.7 cM/loci. This genetic map was also compared with the physical map of diploid progenitors of tetraploid peanut, resulting in an overall co-linearity of about 60% with the average co-linearity of 68% for the A sub-genome and 47% for the B sub-genome. The analysis using the improved genetic map and multi-season (2010-2013) phenotypic data resulted in the identification of 48 quantitative trait loci (QTLs) with phenotypic variance explained (PVE) from 3.88 to 29.14%. Of the 48 QTLs, six QTLs were identified for resistance to TSWV, 22 QTLs for early leaf spot (ELS) and 20 QTLs for late leaf spot (LLS), which included four, six, and six major QTLs (PVE larger than 10%) for each disease, respectively. A total of six major genomic regions (MGR) were found to have QTLs controlling more than one disease resistance. The identified QTLs and resistance gene-rich MGRs will facilitate further discovery of resistance genes and development of molecular markers for these important diseases.

Identification of QTLs associated with oil content and mapping FAD2 genes and their relative contribution to oil quality in peanut (Arachis hypogaea L.).

BACKGROUND: Peanut is one of the major source for human consumption worldwide and its seed contain approximately 50% oil. Improvement of oil content and quality traits (high oleic and low linoleic acid) in peanut could be accelerated by exploiting linked markers through molecular breeding. The objective of this study was to identify QTLs associated with oil content, and estimate relative contribution of FAD2 genes (ahFAD2A and ahFAD2B) to oil quality traits in two recombinant inbred line (RIL) populations. RESULTS: Improved genetic linkage maps were developed for S-population (SunOleic 97R × NC94022) with 206 (1780.6 cM) and T-population (Tifrunner × GT-C20) with 378 (2487.4 cM) marker loci. A total of 6 and 9 QTLs controlling oil content were identified in the S- and T-population, respectively. The contribution of each QTL towards oil content variation ranged from 3.07 to 10.23% in the S-population and from 3.93 to 14.07% in the T-population. The mapping positions for ahFAD2A (A sub-genome) and ahFAD2B (B sub-genome) genes were assigned on a09 and b09 linkage groups. The ahFAD2B gene (26.54%, 25.59% and 41.02% PVE) had higher phenotypic effect on oleic acid (C18:1), linoleic acid (C18:2), and oleic/linoleic acid ratio (O/L ratio) than ahFAD2A gene (8.08%, 6.86% and 3.78% PVE). The FAD2 genes had no effect on oil content. This study identified a total of 78 main-effect QTLs (M-QTLs) with up to 42.33% phenotypic variation (PVE) and 10 epistatic QTLs (E-QTLs) up to 3.31% PVE for oil content and quality traits. CONCLUSIONS: A total of 78 main-effect QTLs (M-QTLs) and 10 E-QTLs have been detected for oil content and oil quality traits. One major QTL (more than 10% PVE) was identified in both the populations for oil content with source alleles from NC94022 and GT-C20 parental genotypes. FAD2 genes showed high effect for oleic acid (C18:1), linoleic acid (C18:2), and O/L ratio while no effect on total oil content. The information on phenotypic effect of FAD2 genes for oleic acid, linoleic acid and O/L ratio, and oil content will be applied in breeding selection.

Oil, fatty acid, flavonoid, and resveratrol content variability and FAD2A functional SNP genotypes in the U.S. peanut mini-core collection.

Peanut seeds contain high amounts of oil and protein as well as some useful bioactive phytochemicals which can contribute to human health. The U.S. peanut mini-core collection is an important genetic resource for improving seed quality and developing new cultivars. Variability of seed chemical composition within the mini-core was evaluated from freshly harvested seeds for two years. Oil, fatty acid composition, and flavonoid/resveratrol content were quantified by NMR, GC, and HPLC, respectively. Significant variability was detected in seed chemical composition among accessions and botanical varieties. Accessions were further genotyped with a functional SNP marker from the FAD2A gene using real-time PCR and classified into three genotypes with significantly different O/L ratios: wild type (G/G with a low O/L ratio <1.7), heterozygote (G/A with O/L ratio >1.4 but <1.7), and mutant (A/A with a high O/L ratio >1.7). The results from real-time PCR genotyping and GC fatty acid analysis were consistent. Accessions with high amounts of oil, quercetin, high seed weight, and O/L ratio were identified. The results from this study may be useful not only for peanut breeders, food processors, and product consumers to select suitable accessions or cultivars but also for curators to potentially expand the mini-core collection.

Evaluation of five peanut (Arachis hypogaea) genotypes to identify drought responsive mechanisms utilising candidate-gene approach.

Drought can significantly limit yield and quality in peanut (Arachis hypogaea L.), depending on its timing, duration and severity. The objective of this study was to identify potential molecular mechanism(s) utilising a candidate-gene approach in five peanut genotypes with contrasting drought responses. An early season drought stress treatment was applied under environmentally controlled rain-out shelters. When water was completely withheld for 3 weeks, no physical differences were observed for treated plants compared with their fully irrigated counterparts as indicated by relative water content; however, yield, grades (total sound mature kernel, TSMK), specific leaf area, and leaf dry matter content showed significant differences. Comparing expression levels of candidate genes, 'C76-16' exhibited significantly higher levels for CuZnSOD, NsLTP and drought protein 1 week earlier compared to the other genotypes, followed by significantly lower levels for the same genes. This suggested an early recognition of drought in C76-16 followed by an acclimation response. Cultivar 'Georgia Green' showed different patterns of gene-expression than C76-16. AP-3, a susceptible genotype, showed generally lower levels of gene-expression than C76-16 and Georgia Green. Myo-inositol phosphate synthase gene-expression showed high levels in irrigated treatment, ranging from 4-fold for 08T-12 to 12-fold for Georgia Green, but were significantly inhibited in drought treatment after 2 weeks of drought and after recovery.