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Key Message: Stable QTL for pod and kernel traits were co-localized on chromosome Arahy05, and an INDEL marker at 106,411,957 on Arahy05 was developed and validated to be useful for marker-assisted selection of kernel weight. Pod and kernel traits, such as hundred pod weight (HPW), and hundred kernel weight (HKW), along with pod and kernel sizes, are pivotal determinants of yield in peanut breeding programs. This study sought to identify quantitative trait loci (QTL) that are associated with these pod and kernel traits in peanuts. To achieve this, a recombinant inbred line (RIL) population, was derived from a cross between Yuhua15, a cultivar known for its high yield, and a germplasm accession W1202. The investigation uncovered stable and major QTL that are significantly associated with both pod and kernel weight and were consistently co-localized on chromosomes Arahy05 and Arahy08. Furthermore, an INDEL marker was identified and characterized in the QTL interval on Arahy05. An extensive re-sequencing analysis comprising 395 germplasm accessions led to the discovery of two principal haplotypes within a 500-kb window flanking the aforementioned INDEL marker. The haplotypes exhibited a significant correlation with the HKW in our diverse panel of germplasm accessions. Notably, the 170 accessions harboring the haplotype associated with an increased HKW primarily represented botanical varieties, specifically Arachis hypogaea var. hypogaea and A. hypogaea var. hirsuta. On the other hand, the 137 accessions associated with the alternative haplotype, which corresponded to a reduced HKW, were predominately identified as belonging to botanical varieties within A. hypogaea subsp. fastigiata. The INDEL marker located on Arahy05, which demonstrates close linkage to the pod and kernel traits, would be an efficient approach for marker-assisted selection (MAS) of pod and kernel weight in breeding programs., (© 2024. The Author(s).)

Cultivated peanut (Arachis hypogaea L.) is a widely grown oilseed crop worldwide; however, the events leading to its origin and diversification are not fully understood. Here by combining chloroplast and whole-genome sequence data from a large germplasm collection, we show that the two subspecies of A. hypogaea (hypogaea and fastigiata) likely arose from distinct allopolyploidization and domestication events. Peanut genetic clusters were then differentiated in relation to dissemination routes and breeding efforts. A combination of linkage mapping and genome-wide association studies allowed us to characterize genes and genomic regions related to main peanut morpho-agronomic traits, namely flowering pattern, inner tegument color, growth habit, pod/seed weight and oil content. Together, our findings shed light on the evolutionary history and phenotypic diversification of peanuts and might be of broad interest to plant breeders., (© 2024. The Author(s).)

For peanut, the lack of stable cytological markers is a barrier to tracking specific chromosomes, elucidating the genetic relationships between genomes and identifying chromosomal variations. Chromosome mapping using single-copy oligonucleotide (oligo) probe libraries has unique advantages for identifying homologous chromosomes and chromosomal rearrangements. In this study, we developed two whole-chromosome single-copy oligo probe libraries, LS-7A and LS-8A, based on the reference genome sequences of chromosomes 7A and 8A of Arachis duranensis . Fluorescence in situ hybridization (FISH) analysis confirmed that the libraries could specifically paint chromosomes 7 and 8. In addition, sequential FISH and electronic localization of LS-7A and LS-8A in A. duranensis ( AA) and A. ipaensis (BB) showed that chromosomes 7A and 8A contained translocations and inversions relative to chromosomes 7B and 8B. Analysis of the chromosomes of wild Arachis species using LS-8A confirmed that this library could accurately and effectively identify A genome species. Finally, LS-7A and LS-8A were used to paint the chromosomes of interspecific hybrids and their progenies, which verified the authenticity of the interspecific hybrids and identified a disomic addition line. This study provides a model for developing specific oligo probes to identify the structural variations of other chromosomes in Arachis and demonstrates the practical utility of LS-7A and LS-8A., Competing Interests: The authors declare that they have no competing interests.

Background: This study aims to decipher the genetic basis governing yield components and quality attributes of peanuts, a critical aspect for advancing molecular breeding techniques. Integrating genotype re-sequencing and phenotypic evaluations of seven yield components and two grain quality traits across four distinct environments allowed for the execution of a genome-wide association study (GWAS)., Results: The nine phenotypic traits were all continuous and followed a normal distribution. The broad heritability ranged from 88.09 to 98.08%, and the genotype-environment interaction effects were all significant. There was a highly significant negative correlation between protein content (PC) and oil content (OC). The 10× genome re-sequencing of 199 peanut accessions yielded a total of 631,988 high-quality single nucleotide polymorphisms (SNPs), with 374 significant SNP loci identified in association with the nine traits of interest. Notably, 66 of these pertinent SNPs were detected in multiple environments, and 48 of them were linked to multiple traits of interest. Five loci situated on chromosome 16 (Chr16) exhibited pleiotropic effects on yield traits, accounting for 17.64-32.61% of the observed phenotypic variation. Two loci on Chr08 were found to be strongly associated with protein and oil contents, accounting for 12.86% and 14.06% of their respective phenotypic variations, respectively. Linkage disequilibrium (LD) block analysis of these seven loci unraveled five nonsynonymous variants, leading to the identification of one yield-related candidate gene and two quality-related candidate genes. The correlation between phenotypic variation and SNP loci in these candidate genes was validated by Kompetitive allele-specific PCR (KASP) marker analysis., Conclusions: Overall, molecular markers were developed for genetic loci associated with yield and quality traits through a GWAS investigation of 199 peanut accessions across four distinct environments. These molecular tools can aid in the development of desirable peanut germplasm with an equilibrium of yield and quality through marker-assisted breeding., (© 2024. The Author(s).)

Background: Bacterial wilt caused by Ralstonia solanacearum severely affects peanut (Arachis hypogaea L.) yields. The breeding of resistant cultivars is an efficient means of controlling plant diseases. Therefore, identification of resistance genes effective against bacterial wilt is a matter of urgency. The lack of a reference genome for a resistant genotype severely hinders the process of identification of resistance genes in peanut. In addition, limited information is available on disease resistance-related pathways in peanut., Results: Full-length transcriptome data were used to generate wilt-resistant and -susceptible transcript pools. In total, 253,869 transcripts were retained to form a reference transcriptome for RNA-sequencing data analysis. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of differentially expressed genes revealed the plant-pathogen interaction pathway to be the main resistance-related pathway for peanut to prevent bacterial invasion and calcium plays an important role in this pathway. Glutathione metabolism was enriched in wilt-susceptible genotypes, which would promote glutathione synthesis in the early stages of pathogen invasion. Based on our previous quantitative trait locus (QTL) mapping results, the genes arahy.V6I7WA and arahy.MXY2PU, which encode nucleotide-binding site-leucine-rich repeat receptor proteins, were indicated to be associated with resistance to bacterial wilt., Conclusions: This study identified several pathways associated with resistance to bacterial wilt and identified candidate genes for bacterial wilt resistance in a major QTL region. These findings lay a foundation for investigation of the mechanism of resistance to bacterial wilt in peanut., (© 2024. The Author(s).)

Phenylalanine ammonia-lyase (PAL) is an essential enzyme in the phenylpropanoid pathway, in which numerous aromatic intermediate metabolites play significant roles in plant growth, adaptation, and disease resistance. Cultivated peanuts are highly susceptible to Aspergillus flavus L. infection. Although PAL genes have been characterized in various major crops, no systematic studies have been conducted in cultivated peanuts, especially in response to A. flavus infection. In the present study, a systematic genome-wide analysis was conducted to identify PAL genes in the Arachis hypogaea L. genome. Ten AhPAL genes were distributed unevenly on nine A. hypogaea chromosomes. Based on phylogenetic analysis, the AhPAL proteins were classified into three groups. Structural and conserved motif analysis of PAL genes in A. hypogaea revealed that all peanut PAL genes contained one intron and ten motifs in the conserved domains. Furthermore, synteny analysis indicated that the ten AhPAL genes could be categorized into five pairs and that each AhPAL gene had a homologous gene in the wild-type peanut. Cis-element analysis revealed that the promoter region of the AhPAL gene family was rich in stress- and hormone-related elements. Expression analysis indicated that genes from Group I ( AhPAL1 and AhPAL2 ), which had large number of ABRE, WUN, and ARE elements in the promoter, played a strong role in response to A. flavus stress.

The capability of embryogenic callus induction is a prerequisite for in vitro plant regeneration. However, embryogenic callus induction is strongly genotype-dependent, thus hindering the development of in vitro plant genetic engineering technology. In this study, to examine the genetic variation in embryogenic callus induction rate (CIR) in peanut ( Arachis hypogaea L.) at the seventh, eighth, and ninth subcultures (T7, T8, and T9, respectively), we performed genome-wide association studies (GWAS) for CIR in a population of 353 peanut accessions. The coefficient of variation of CIR among the genotypes was high in the T7, T8, and T9 subcultures (33.06%, 34.18%, and 35.54%, respectively), and the average CIR ranged from 1.58 to 1.66. A total of 53 significant single-nucleotide polymorphisms (SNPs) were detected (based on the threshold value -log 10 ( p ) = 4.5). Among these SNPs, SNPB03-83801701 showed high phenotypic variance and neared a gene that encodes a peroxisomal ABC transporter 1. SNPA05-94095749, representing a nonsynonymous mutation, was located in the Arahy.MIX90M locus (encoding an auxin response factor 19 protein) at T8, which was associated with callus formation. These results provide guidance for future elucidation of the regulatory mechanism of embryogenic callus induction in peanut.

Background: Pod shell thickness (PST) is an important agronomic trait of peanut because it affects the ability of shells to resist pest infestations and pathogen attacks, while also influencing the peanut shelling process. However, very few studies have explored the genetic basis of PST., Results: An F 2 segregating population derived from a cross between the thick-shelled cultivar Yueyou 18 (YY18) and the thin-shelled cultivar Weihua 8 (WH8) was used to identify the quantitative trait loci (QTLs) for PST. On the basis of a bulked segregant analysis sequencing (BSA-seq), four QTLs were preliminarily mapped to chromosomes 3, 8, 13, and 18. Using the genome resequencing data of YY18 and WH8, 22 kompetitive allele-specific PCR (KASP) markers were designed for the genotyping of the F 2 population. Two major QTLs (qPSTA08 and qPSTA18) were identified and finely mapped, with qPSTA08 detected on chromosome 8 (0.69-Mb physical genomic region) and qPSTA18 detected on chromosome 18 (0.15-Mb physical genomic region). Moreover, qPSTA08 and qPSTA18 explained 31.1-32.3% and 16.7-16.8% of the phenotypic variation, respectively. Fifteen genes were detected in the two candidate regions, including three genes with nonsynonymous mutations in the exon region. Two molecular markers (Tif2_A08_31713024 and Tif2_A18_7198124) that were developed for the two major QTL regions effectively distinguished between thick-shelled and thin-shelled materials. Subsequently, the two markers were validated in four F 2:3 lines selected., Conclusions: The QTLs identified and molecular markers developed in this study may lay the foundation for breeding cultivars with a shell thickness suitable for mechanized peanut shelling., (© 2024. The Author(s).)

Peanut (Arachis hypogaea L.) is an important oilseed crop worldwide. Improving its yield is crucial for sustainable peanut production to meet increasing food and industrial requirements. Deciphering the genetic control underlying peanut kernel weight and size, which are essential components of peanut yield, would facilitate high-yield breeding. A high-density single nucleotide polymorphism (SNP)-based linkage map was constructed using a recombinant inbred lines (RIL) population derived from a cross between the variety Yuanza9102 and a germplasm accession wt09-0023. Kernel weight and size quantitative trait loci (QTLs) were co-localized to a 0.16 Mb interval on Arahy07 using inclusive composite interval mapping (ICIM). Analysis of SNP, and Insertion or Deletion (INDEL) markers in the QTL interval revealed a gene encoding a pentatricopeptide repeat (PPR) superfamily protein as a candidate closely linked with kernel weight and size in cultivated peanut. Examination of the PPR gene family indicated a high degree of collinearity of PPR genes between A. hypogaea and its diploid progenitors, Arachis duranensis and Arachis ipaensis. The candidate PPR gene, Arahy.JX1V6X, displayed a constitutive expression pattern in developing seeds. These findings lay a foundation for further fine mapping of QTLs related to kernel weight and size, as well as validation of candidate genes in cultivated peanut., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Key Message: QTLs for growth habit are identified on Arahy.15 and Arahy.06 in peanut, and diagnostic markers are developed and validated for further use in marker-assisted breeding. Peanut is a unique legume crop because its pods develop and mature underground. The pegs derive from flowers following pollination, then reach the ground and develop into pods in the soil. Pod number per plant is influenced by peanut growth habit (GH) that has been categorized into four types, including erect, bunch, spreading and prostrate. Restricting pod development at the plant base, as would be the case for peanut plants with upright lateral branches, would decrease pod yield. On the other hand, GH characterized by spreading lateral branches on the ground would facilitate pod formation on the nodes, thereby increasing yield potential. We describe herein an investigation into the GH traits of 521 peanut recombinant inbred lines grown in three distinct environments. Quantitative trait loci (QTLs) for GH were identified on linkage group (LG) 15 between 203.1 and 204.2 cM and on LG 16 from 139.1 to 139.3 cM. Analysis of resequencing data in the identified QTL regions revealed that single nucleotide polymorphism (SNP) or insertion and/or deletion (INDEL) at Arahy15.156854742, Arahy15.156931574, Arahy15.156976352 and Arahy06.111973258 may affect the functions of their respective candidate genes, Arahy.QV02Z8, Arahy.509QUQ, Arahy.ATH5WE and Arahy.SC7TJM. These SNPs and INDELs in relation to peanut GH were further developed for KASP genotyping and tested on a panel of 77 peanut accessions with distinct GH features. This study validates four diagnostic markers that may be used to distinguish erect/bunch peanuts from spreading/prostrate peanuts, thereby facilitating marker-assisted selection for GH traits in peanut breeding., (© 2023. The Author(s).)

Systemic acquired resistance is an essential immune response that triggers a broad-spectrum disease resistance throughout the plant. In the present study, we identified a peanut lesion mimic mutant m14 derived from an ethyl methane sulfonate-mutagenized mutant pool of peanut cultivar "Yuanza9102." Brown lesions were observed in the leaves of an m14 mutant from seedling stage to maturity. Using MutMap together with bulked segregation RNA analysis approaches, a G-to-A point mutation was identified in the exon region of candidate gene Arahy.R60CUW, which is the homolog of AtNPR3 (Nonexpresser of PR genes) in Arabidopsis. This point mutation caused a transition from Gly to Arg within the C-terminal transactivation domain of AhNPR3A. The mutation of AhNPR3A showed no effect in the induction of PR genes when treated with salicylic acid. Instead, the mutation resulted in upregulation of WRKY genes and several PR genes, including pathogenesis-related thaumatin- and chitinase-encoding genes, which is consistent with the resistant phenotype of m14 to leaf spot disease. Further study on the AhNPR3A gene will provide valuable insights into understanding the molecular mechanism of systemic acquired resistance in peanut. Moreover, our results indicated that a combination of MutMap and bulked segregation RNA analysis is an effective method for identifying genes from peanut mutants., (© 2022 Society for Experimental Biology and John Wiley & Sons Ltd.)

Key Message: A major QTL, qBWA12, was fine mapped to a 216.68 kb physical region, and A12.4097252 was identified as a useful KASP marker for breeding peanut varieties resistant to bacterial wilt. Bacterial wilt, caused by Ralstonia solanacearum, is a major disease detrimental to peanut production in China. Breeding disease-resistant peanut varieties is the most economical and effective way to prevent the disease and yield loss. Fine mapping the QTLs for bacterial wilt resistance is critical for the marker-assisted breeding of disease-resistant varieties. A recombinant inbred population comprising 521 lines was used to construct a high-density genetic linkage map and to identify QTLs for bacterial wilt resistance following restriction-site-associated DNA sequencing. The genetic map, which included 5120 SNP markers, covered a length of 3179 cM with an average marker distance of 0.6 cM. Four QTLs for bacterial wilt resistance were mapped on four chromosomes. One major QTL, qBWA12, with LOD score of 32.8-66.0 and PVE of 31.2-44.8%, was stably detected in all four development stages investigated over the 3 trial years. Additionally, qBWA12 spanned a 2.7 cM region, corresponding to approximately 0.4 Mb and was fine mapped to a 216.7 kb region by applying KASP markers that were polymorphic between the two parents based on whole-genome resequencing data. In a large collection of breeding and germplasm lines, it was proved that KASP marker A12.4097252 can be applied for the marker-assisted breeding to develop peanut varieties resistant to bacterial wilt. Of the 19 candidate genes in the region covered by qBWA12, nine NBS-LRR genes should be further investigated regarding their potential contribution to the resistance of peanut against bacterial wilt., (© 2022. The Author(s).)

Pod size is an important yield-influencing trait in peanuts. It is affected by plant hormones and identifying the genes related to these hormones may contribute to pod-related trait improvements in peanut breeding programs. However, there is limited information on the molecular mechanisms of plant hormones that regulate pod size in peanuts. We identified a mutant with an extremely small pod ( spm ) from Yuanza 9102 (WT) by 60 Co γ-radiation mutagenesis. The length and width of the natural mature pod in spm were only 71.34% and 73.36% of those in WT, respectively. We performed comparative analyses for morphological characteristics, anatomy, physiology, and global transcriptome between spm and WT pods. Samples were collected at 10, 20, and 30 days after peg elongation into the soil, representing stages S1, S2, and S3, respectively. The differences in pod size between WT and spm were seen at stage S1 and became even more striking at stages S2 and S3. The cell sizes of the pods were significantly smaller in spm than in WT at stages S1, S2, and S3. These results suggested that reduced cell size may be one of the important contributors for the small pod in spm . The contents of indole-3-acetic acid (IAA), gibberellin (GA), and brassinosteroid (BR) were also significantly lower in spm pods than those in WT pods at all three stages. RNA-Seq analyses showed that 1,373, 8,053, and 3,358 differently expressed genes (DEGs) were identified at stages S1, S2, and S3, respectively. Functional analyses revealed that a set of DEGs was related to plant hormone biosynthesis, plant hormone signal transduction pathway, and cell wall biosynthesis and metabolism. Furthermore, several hub genes associated with plant hormone biosynthesis and signal transduction pathways were identified through weighted gene co-expression network analysis. Our results revealed that IAA, GA, and BR may be important regulators in controlling pod size by regulating cell size in peanuts. This study provides helpful information for the understanding of the complex mechanisms of plant hormones in controlling pod size by regulating the cell size in peanuts and will facilitate the improvement of peanut breeding., Competing Interests: The authors declare there are no competing interests., (©2022 Wang et al.)

Background: Chromosomal variants play important roles in crop breeding and genetic research. The development of single-stranded oligonucleotide (oligo) probes simplifies the process of fluorescence in situ hybridization (FISH) and facilitates chromosomal identification in many species. Genome sequencing provides rich resources for the development of oligo probes. However, little progress has been made in peanut due to the lack of efficient chromosomal markers. Until now, the identification of chromosomal variants in peanut has remained a challenge., Results: A total of 114 new oligo probes were developed based on the genome-wide tandem repeats (TRs) identified from the reference sequences of the peanut variety Tifrunner (AABB, 2n = 4x = 40) and the diploid species Arachis ipaensis (BB, 2n = 2x = 20). These oligo probes were classified into 28 types based on their positions and overlapping signals in chromosomes. For each type, a representative oligo was selected and modified with green fluorescein 6-carboxyfluorescein (FAM) or red fluorescein 6-carboxytetramethylrhodamine (TAMRA). Two cocktails, Multiplex #3 and Multiplex #4, were developed by pooling the fluorophore conjugated probes. Multiplex #3 included FAM-modified oligo TIF-439, oligo TIF-185-1, oligo TIF-134-3 and oligo TIF-165. Multiplex #4 included TAMRA-modified oligo Ipa-1162, oligo Ipa-1137, oligo DP-1 and oligo DP-5. Each cocktail enabled the establishment of a genome map-based karyotype after sequential FISH/genomic in situ hybridization (GISH) and in silico mapping. Furthermore, we identified 14 chromosomal variants of the peanut induced by radiation exposure. A total of 28 representative probes were further chromosomally mapped onto the new karyotype. Among the probes, eight were mapped in the secondary constrictions, intercalary and terminal regions; four were B genome-specific; one was chromosome-specific; and the remaining 15 were extensively mapped in the pericentric regions of the chromosomes., Conclusions: The development of new oligo probes provides an effective set of tools which can be used to distinguish the various chromosomes of the peanut. Physical mapping by FISH reveals the genomic organization of repetitive oligos in peanut chromosomes. A genome map-based karyotype was established and used for the identification of chromosome variations in peanut following comparisons with their reference sequence positions.

Background: Web blotch is one of the most important foliar diseases worldwide in peanut (Arachis hypogaea L.). The identification of quantitative trait loci (QTLs) for peanut web blotch resistance represents the basis for gene mining and the application of molecular breeding technologies., Results: In this study, a peanut recombinant inbred line (RIL) population was used to map QTLs for web blotch resistance based on high-throughput genome-wide sequencing. Frequency distributions of disease grade and disease index in five environments indicated wide phenotypic variations in response to web blotch among RILs. A high-density genetic map was constructed, containing 3634 bin markers distributed on 20 peanut linkage groups (LGs) with an average genetic distance of 0.5 cM. In total, eight QTLs were detected for peanut web blotch resistance in at least two environments, explaining from 2.8 to 15.1% of phenotypic variance. Two major QTLs qWBRA04 and qWBRA14 were detected in all five environments and were linked to 40 candidate genes encoding nucleotide-binding site leucine-rich repeat (NBS-LRR) or other proteins related to disease resistances., Conclusions: The results of this study provide a basis for breeding peanut cultivars with web blotch resistance.

Background: Arachis contains 80 species that carry many beneficial genes that can be utilized in the genetic improvement of peanut (Arachis hypogaea L. 2n = 4x = 40, genome AABB). Chromosome engineering is a powerful technique by which these genes can be transferred and utilized in cultivated peanut. However, their small chromosomes and insufficient cytological markers have made chromosome identification and studies relating to genome evolution quite difficult. The development of efficient cytological markers or probes is very necessary for both chromosome engineering and genome discrimination in cultivated peanut., Results: A simple and efficient oligonucleotide multiplex probe to distinguish genomes, chromosomes, and chromosomal aberrations of peanut was developed based on eight single-stranded oligonucleotides (SSONs) derived from repetitive sequences. High-resolution karyotypes of 16 Arachis species, two interspecific F 1 hybrids, and one radiation-induced M 1 plant were then developed by fluorescence in situ hybridization (FISH) using oligonucleotide multiplex, 45S and 5S rDNAs, and genomic in situ hybridization (GISH) using total genomic DNA of A. duranensis (2n = 2x = 20, AA) and A. ipaënsis (2n = 2x = 20, BB) as probes. Genomes, chromosomes, and aberrations were clearly identifiable in the established karyotypes. All eight cultivars had similar karyotypes, whereas the eight wild species exhibited various chromosomal variations. In addition, a chromosome-specific SSON library was developed based on the single-copy sequence of chromosome 6A of A. duranensis. In combination with repetitive SSONs and rDNA FISH, the single-copy SSON library was applied to identify the corresponding A3 chromosome in the A. duranensis karyotype., Conclusions: The development of repetitive and single-copy SSON probes for FISH and GISH provides useful tools for the differentiation of chromosomes and identification of structural chromosomal rearrangement. It facilitates the development of high-resolution karyotypes and detection of chromosomal variations in Arachis species. To our knowledge, the methodology presented in this study demonstrates for the first time the correlation between a sequenced chromosome region and a cytologically identified chromosome in peanut.

Cultivated peanut (Arachis hypogaea L.) were classified into six botanical varieties according to the morphological characteristics. However, their genetic evolutionary relationships at the genome-wide level were still unclear. A total of 320 peanut accessions, including four of the six botanical varieties, and 37,128 high-quality single nucleotide polymorphisms (SNPs) detected by tunable genotyping-by-sequencing (tGBS) were used to reveal the evolutionary relationships among different botanical varieties and verify the phenotypic classification. A phylogenetic tree indicated that the tested accessions were grouped into three clusters. Almost all of the peanut accessions in cluster C1 belong to var. fastigiata, and clusters C2 and C3 mainly consisted of accessions from var. vulgaris and subsp. hypogaea, respectively. The results of a principal component analysis were consistent with relationships revealed in the phylogenetic tree. Population structure analysis showed that var. fastigiata and var. vulgaris were not separated when K = 2 (subgroup number), whereas they were clearly divided when K = 3. However, var. hypogaea and var. hirsuta could not be distinguished from each other all the way. The nucleotide diversity (π) value implied that var. vulgaris exhibited the highest genetic diversity (0.048), followed by var. fastigiata (0.035) and subsp. hypogaea (0.012), which is consistent with the result of phylogenetic tree. Moreover, the fixation index (F ST ) value confirmed that var. fastigiata and var. vulgaris were closely related to each other (F ST  = 0.284), while both of them were clearly distinct from var. hypogaea (F ST  > 0.4). The present study confirmed the traditional botanical classifications of cultivated peanut at the genome-wide level. Furthermore, the reliable SNPs identified in this study may be a valuable resource for peanut breeders.

Late leaf spot (LLS) is a major foliar disease in peanut (A. hypogaea L.) worldwide, causing significant losses of potential yield in the absence of fungicide applications. Mutants are important materials to study the function of disease-related genes. In this study, the mutant line M14 was derived from cultivar Yuanza 9102 treated with EMS. Yuanza 9102 was selected from an interspecific cross of cultivar Baisha 1016 with A. diogoi, and is resistant to several fungal diseases. By contrast, the M14 was highly susceptible to late leaf spot. RNA-Seq analysis in the leaf tissues of the M14 and its wild type Yuanza 9102 under pathogen challenge showed 2219 differentially expressed genes including1317 up-regulated genes and 902 down-regulated genes. Of these genes, 1541, 1988, 1344, 643 and 533 unigenes were obtained and annotated by public protein databases of SwissPort, TrEMBL, gene ontology (GO), KEGG and clusters of orthologous groups (COG), respectively. Differentially expressed genes (DEGs) showed that expression of inducible pathogenesis-related (PR) proteins was significantly up-regulated; in the meantime DEGs related to photosynthesis were down-regulated in the susceptible M14 in comparison to the resistant WT. Moreover, the up-regulated WRKY transcription factors and down-regulated plant hormones related to plant growth were detected in the M14. The results suggest that down-regulated chloroplast genes, up-regulated WRKY transcription factors, and depressed plant hormones related to plant growth in the M14 might coordinately render the susceptibility though there was a significant high level of PRs. Those negative effectors might be triggered in the susceptible plant by fungal infection and resulted in reduction of photosynthesis and phytohormones and led to symptom formation.

Cultivated peanut is grown worldwide as rich-source of oil and protein. A broad genetic base is needed for cultivar improvement. The objectives of this study were to develop highly informative simple sequence repeat (SSR) markers and to assess the genetic diversity and population structure of peanut cultivars and breeding lines from different breeding programs in China, India and the US. A total of 111 SSR markers were selected for this study, resulting in a total of 472 alleles. The mean values of gene diversity and polymorphic information content (PIC) were 0.480 and 0.429, respectively. Country-wise analysis revealed that alleles per locus in three countries were similar. The mean gene diversity in the US, China and India was 0.363, 0.489 and 0.47 with an average PIC of 0.323, 0.43 and 0.412, respectively. Genetic analysis using the STRUCTURE divided these peanut lines into two populations (P1, P2), which was consistent with the dendrogram based on genetic distance (G1, G2) and the clustering of principal component analysis. The groupings were related to peanut market types and the geographic origin with a few admixtures. The results could be used by breeding programs to assess the genetic diversity of breeding materials to broaden the genetic base and for molecular genetics studies., (Published 2015. This article is a U.S. Government work and is in the public domain in the USA.)

Key message: QTLs for growth habit are identified on Arahy.15 and Arahy.06 in peanut, and diagnostic markers are developed and validated for further use in marker-assisted breeding. Peanut is a unique legume crop because its pods develop and mature underground. The pegs derive from flowers following pollination, then reach the ground and develop into pods in the soil. Pod number per plant is influenced by peanut growth habit (GH) that has been categorized into four types, including erect, bunch, spreading and prostrate. Restricting pod development at the plant base, as would be the case for peanut plants with upright lateral branches, would decrease pod yield. On the other hand, GH characterized by spreading lateral branches on the ground would facilitate pod formation on the nodes, thereby increasing yield potential. We describe herein an investigation into the GH traits of 521 peanut recombinant inbred lines grown in three distinct environments. Quantitative trait loci (QTLs) for GH were identified on linkage group (LG) 15 between 203.1 and 204.2 cM and on LG 16 from 139.1 to 139.3 cM. Analysis of resequencing data in the identified QTL regions revealed that single nucleotide polymorphism (SNP) or insertion and/or deletion (INDEL) at Arahy15.156854742, Arahy15.156931574, Arahy15.156976352 and Arahy06.111973258 may affect the functions of their respective candidate genes, Arahy.QV02Z8, Arahy.509QUQ, Arahy.ATH5WE and Arahy.SC7TJM. These SNPs and INDELs in relation to peanut GH were further developed for KASP genotyping and tested on a panel of 77 peanut accessions with distinct GH features. This study validates four diagnostic markers that may be used to distinguish erect/bunch peanuts from spreading/prostrate peanuts, thereby facilitating marker-assisted selection for GH traits in peanut breeding. [ABSTRACT FROM AUTHOR]

Peanut (Arachis hypogaea) is an important economic crop that is susceptible to various foliar diseases. Web blotch, which is caused by Phoma arachidicola, is one of the most serious peanut foliar diseases because it affects the leaves and results in substantial yield losses. In this study, the P. arachidicola infection process and the response of peanut plants to infection were revealed through a cytological study of resistant and susceptible peanut varieties. Conidial germination rates, appressorium formation and resistance responses, including papillae formation and the hypersensitive response (HR), were analysed at different infection stages. Notably, the basal infection response of susceptible varieties resulted in limited papillae formation and a single HR, which was followed by programmed cell death. The papillae formation frequency and HR frequency were significantly greater in the resistant varieties than in the susceptible varieties at 84 h postinoculation. The primary difference between the two resistant varieties was the superior HR of Zheng8903 compared with that of Yuhua15. The complex amphidiploid peanut genome may explain the complexity and diversity of the resistance responses. The interactions of functional orthologs should be clarified and perhaps modulated in future investigations. The results of this study revealed the differences in the resistance responses among peanut germplasm and may be relevant for the breeding of new peanut varieties highly resistant to P. arachidicola. [ABSTRACT FROM AUTHOR]

Cultivated peanut (Arachis hypogaea L.), a cosmopolitan oil crop, is susceptible to a variety of pathogens, especially Aspergillus flavus L., which not only vastly reduce the quality of peanut products but also seriously threaten food safety for the contamination of aflatoxin. However, the key genes related to resistance to Aspergillus flavus L. in peanuts remain unclear. This study identifies hub genes positively associated with resistance to A. flavus in two genotypes by comparative transcriptome and weighted gene co-expression network analysis (WGCNA) method. Compared with susceptible genotype (Zhonghua 12, S), the rapid response to A. flavus and quick preparation for the translation of resistance-related genes in the resistant genotype (J-11, R) may be the drivers of its high resistance. WGCNA analysis revealed that 18 genes encoding pathogenesis-related proteins (PR10), 1-aminocyclopropane-1-carboxylate oxidase (ACO1), MAPK kinase, serine/threonine kinase (STK), pattern recognition receptors (PRRs), cytochrome P450, SNARE protein SYP121, pectinesterase, phosphatidylinositol transfer protein, and pentatricopeptide repeat (PPR) protein play major and active roles in peanut resistance to A. flavus. Collectively, this study provides new insight into resistance to A. flavus by employing WGCNA, and the identification of hub resistance-responsive genes may contribute to the development of resistant cultivars by molecular-assisted breeding. [ABSTRACT FROM AUTHOR]

Key message: A major QTL, qBWA12, was fine mapped to a 216.68 kb physical region, and A12.4097252 was identified as a useful KASP marker for breeding peanut varieties resistant to bacterial wilt. Bacterial wilt, caused by Ralstonia solanacearum, is a major disease detrimental to peanut production in China. Breeding disease-resistant peanut varieties is the most economical and effective way to prevent the disease and yield loss. Fine mapping the QTLs for bacterial wilt resistance is critical for the marker-assisted breeding of disease-resistant varieties. A recombinant inbred population comprising 521 lines was used to construct a high-density genetic linkage map and to identify QTLs for bacterial wilt resistance following restriction-site-associated DNA sequencing. The genetic map, which included 5120 SNP markers, covered a length of 3179 cM with an average marker distance of 0.6 cM. Four QTLs for bacterial wilt resistance were mapped on four chromosomes. One major QTL, qBWA12, with LOD score of 32.8–66.0 and PVE of 31.2–44.8%, was stably detected in all four development stages investigated over the 3 trial years. Additionally, qBWA12 spanned a 2.7 cM region, corresponding to approximately 0.4 Mb and was fine mapped to a 216.7 kb region by applying KASP markers that were polymorphic between the two parents based on whole-genome resequencing data. In a large collection of breeding and germplasm lines, it was proved that KASP marker A12.4097252 can be applied for the marker-assisted breeding to develop peanut varieties resistant to bacterial wilt. Of the 19 candidate genes in the region covered by qBWA12, nine NBS-LRR genes should be further investigated regarding their potential contribution to the resistance of peanut against bacterial wilt. [ABSTRACT FROM AUTHOR]

Peanuts (Arachis hypogaea L.) offer numerous healthy benefits, and the production of peanuts has a prominent role in global food security. As a result, it is in the interest of society to improve the productivity and quality of peanuts with transgenic means. However, the lack of a robust phylogeny of cultivated and wild peanut species has limited the utilization of genetic resources in peanut molecular breeding. In this study, a total of 33 complete peanut plastomes were sequenced, analyzed and used for phylogenetic analyses. Our results suggest that sect. Arachis can be subdivided into two lineages. All the cultivated species are contained in Lineage I with AABB and AA are the two predominant genome types present, while species in Lineage II possess diverse genome types, including BB, KK, GG, etc. Phylogenetic studies also indicate that all allotetraploid cultivated peanut species have been derived from a possible maternal hybridization event with one of the diploid Arachis duranensis accessions being a potential AA sub-genome ancestor. In addition, Arachis monticola , a tetraploid wild species, is placed in the same group with all the cultivated peanuts, and it may represent a transitional species, which has been through the recent hybridization event. This research could facilitate a better understanding of the taxonomic status of various Arachis species/accessions and the evolutionary relationship among them, and assists in the correct and efficient use of germplasm resources in breeding efforts to improve peanuts for the benefit of human beings. [ABSTRACT FROM AUTHOR]

APETALA2/ethylene response element-binding factor (AP2/ERF) transcription factors (TFs) have been found to regulate plant growth and development and response to various abiotic stresses. However, detailed information of AP2/ERF genes in peanut against drought has not yet been performed. Herein, 185 AP2/ERF TF members were identified from the cultivated peanut (A. hypogaea cv. Tifrunner) genome, clustered into five subfamilies: AP2 (APETALA2), ERF (ethylene-responsive-element-binding), DREB (dehydration-responsive-element-binding), RAV (related to ABI3/VP), and Soloist (few unclassified factors)). Subsequently, the phylogenetic relationship, intron–exon structure, and chromosomal location of AhAP2/ERF were further characterized. All of these AhAP2/ERF genes were distributed unevenly across the 20 chromosomes, and 14 tandem and 85 segmental duplicated gene pairs were identified which originated from ancient duplication events. Gene evolution analysis showed that A. hypogaea cv. Tifrunner were separated 64.07 and 66.44 Mya from Medicago truncatula L. and Glycine max L., respectively. Promoter analysis discovered many cis -acting elements related to light, hormones, tissues, and stress responsiveness process. The protein interaction network predicted the exitance of functional interaction among families or subgroups. Expression profiles showed that genes from AP2 , ERF , and dehydration-responsive-element-binding subfamilies were significantly upregulated under drought stress conditions. Our study laid a foundation and provided a panel of candidate AP2/ERF TFs for further functional validation to uplift breeding programs of drought-resistant peanut cultivars. [ABSTRACT FROM AUTHOR]

Background: Development of oligonucleotide probes facilitates chromosome identification via fluorescence in situ hybridization (FISH) in many organisms. Results: We report a high throughput and economical method of chromosome identification based on the development of a dye solution containing 2 × saline-sodium citrate (SSC) and oligonucleotide probes. Based on the concentration, staining time, and sequence effects of oligonucleotides, an efficient probe dye of peanut was developed for chromosome identification. To validate the effects of this solution, 200 slides derived from 21 accessions of the cultivated peanut and 30 wild Arachis species were painted to identify Arachis genomes and establish karyotypes. The results showed that one jar of dye could be used to paint 10 chromosome preparations and recycled at least 10 times to efficiently dye more than 100 slides. The A, B, K, F, E, and H genomes showed unique staining karyotype patterns and signal colors. Conclusions: Based on the karyotype patterns of Arachis genomes, we revealed the relationships among the A, B, K, F, E, and H genomes in genus Arachis, and demonstrated the potential for adoption of this oligonucleotide dye solution in practice. [ABSTRACT FROM AUTHOR]