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Plant cell development is an asynchronous process that is governed by multiple layers of gene regulation. However, the correlation between transcriptome and chromatin regulatory events in an allotetraploid species at the single-cell resolution has not been widely studied. Herein, we employed fluorescence-activated nuclei sorting to isolate single nuclei and simultaneously investigate the transcriptome (snRNA-seq) and chromatin accessibility (snATAC-seq) landscapes in the same leaf single-cell of Arachis hypogaea. A total of 5,930 cells with 10,793 expressed genes were classified into 17 cell-clusters and 5,315 chromatin fragments were enriched to target 26,083 genes in the snATAC-seq landscape. The developmental trajectory revealed a conserved ethylene-AP2 module in leaf cell differentiation and provided novel insight for mesophyll and vascular cell development. Additionally, dual-omics described the epidermal progenitor cell development trajectory, primordium and palisade cells were able to convert into spongy cells, and bundle sheath cells developed earlier than other vascular-cells. Further cell-cycle analysis demonstrated that cytokinin biosynthesis promotes mesophyll cell genome replication and lipid pathway participates in guard cell development. snRNA-seq identified that the AT-hook transcription factor AhAHL11promotes leaf area growth by modulating auxin content, but snATAC-seq identified AhBHLH143 displaying contrasting results by repressing leaf development via the jasmonic acid pathway in ectopically expressed Arabidopsis. Conclusively, our study demonstrates that snRNA-seq combined with snATAC-seq is an effective platform for exploring the association between chromatin regulatory events and transcriptional activity across diverse cell types in peanut leaves. The broad application of this approach will enable significant advances in the functional research of tissue growth and development in plant species. Plant cell development is an asynchronous process that is governed by multiple layers of gene regulation. However, the correlation between transcriptome and chromatin regulatory events in an allotetraploid species at the single-cell resolution has not been widely studied. Herein, we employed fluorescence-activated nuclei sorting to isolate single nuclei and simultaneously investigate the transcriptome (snRNA-seq) and chromatin accessibility (snATAC-seq) landscapes in the same leaf single-cell of peanut. Totally 5,930 cells with 10,793 expressed genes were classified into 17 cell-clusters and 5,315 chromatin fragments were enriched to target 26,083 genes in the snATAC-seq landscape. Developmental trajectory revealed a conserved ethylene-AP2 module in leaf cell differentiation and provided novel insights for mesophyll and vascular cells development. Further ell-cycle demonstrated that cytokinin promotes mesophyll-cell genome replication and lipid pathway participates in guard cell development. snRNA-seq identified AhAHL11 promotes leaf area growth by modulating auxin content, but snATAC-seq identified AhBHLH143 repressing leaf development via jasmonic acid pathway. Conclusively, snRNA-seq combined with snATAC-seq is an effective platform for exploring the association between chromatin regulatory events and transcriptional activity across diverse cell-types. The broad application of this approach will enable significant advances in the functional research of tissue growth and development in plant species.

The 14-3-3 protein is a kind of evolutionary ubiquitous protein family highly conserved in eukaryotes. Initially, 14-3-3 proteins were reported in mammalian nervous tissues, but in the last decade, their role in various metabolic pathways in plants established the importance of 14-3-3 proteins. In present study, a total of 22 14-3-3 genes also called as general regulatory factors (GRF) were identified in the peanut (Arachis hypogaea) genome out of which 12 belonged to the ε group whereas 10 of them belonged to the non- ε-group. Tissue-specific expression of identified 14-3-3 genes was studied using transcriptome analysis. The peanut AhGRFi gene was cloned and transformed in Arabidopsis thaliana. The investigation of subcellular localization indicated that AhGRFi is localized in the cytoplasm. Overexpression of the AhGRFi gene in transgenic Arabidopsis shown that under exogenous 1-naphthaleneacetic acid (NAA) treatment, root growth of transgenic plants was inhibited when compared with control. NAA-responsive endogenous relative gene expressions in AhGRFi transgenic Arabidopsis were validated by real-time PCR. The AhGRFi may be involved in the auxin signaling during seedling root development. An in-depth study on the molecular mechanism of this process remains to be further explored.

Summary Single‐cell RNA‐seq (scRNA‐seq) has been highlighted as a powerful tool for the description of human cell transcriptome, but the technology has not been broadly applied in plant cells. Herein, we describe the successful development of a robust protoplast cell isolation system in the peanut leaf. A total of 6,815 single cells were divided into eight cell clusters based on reported marker genes by applying scRNA‐seq. Further, a pseudo‐time analysis was used to describe the developmental trajectory and interaction network of transcription factors (TFs) of distinct cell types during leaf growth. The trajectory enabled re‐investigation of the primordium‐driven development processes of the mesophyll and epidermis. These results suggest that palisade cells likely differentiate into spongy cells, while the epidermal cells originated earlier than the primordium. Subsequently, the developed method integrated multiple technologies to efficiently validate the scRNA‐seq result in a homogenous cell population. The expression levels of several TFs were strongly correlated with epidermal ontogeny in accordance with obtained scRNA‐seq values. Additionally, peanut AHL23 (AT‐HOOK MOTIF NUCLEAR LOCALIZED PROTEIN 23), which is localized in nucleus, promoted leaf growth when ectopically expressed in Arabidopsis by modulating the phytohormone pathway. Together, our study displays that application of scRNA‐seq can provide new hypotheses regarding cell differentiation in the leaf blade of Arachis hypogaea. We believe that this approach will enable significant advances in the functional study of leaf blade cells in the allotetraploid peanut and other plant species.

Plant cell proliferation associated with multiple layers of gene regulation, including modulation of transcriptome by changes in chromatin accessibility. However, cell proliferation is an asynchronous process precluding a temporal understanding of regulatory events leading to single-cell fate commitment. Here, a robust single nucleus RNA sequencing approach, where single nucleus employed for simultaneous investigation of transcriptome (snRNA-seq) and chromatin accessibility (snATAC-seq) landscapes in the same single-cell of Arachis hypogaea leaves. A total of 5,930 leaf cells with 10,793 expressed genes were used to construct development trajectory and characterized large-scale critical differentially expressed genes (DEGs). Additionally, uncovered extending insights of chromatin opening guided 5,315 DEGs expression involved biological pathway determines differentiation direction in distinct cell-types. But obtained members in each cell-clusters not exhibits obvious difference in distinct cell-cycling regulated genome duplication phases. Furthermore, snRNA-seq identified AT-hook transcription factor AhAHL11 promotes leaf area growth by modulating auxin content, but snATAC-seq identified AhBHLH143 displays contrasting results to repress the leaf development by jasmonic acid pathway in ectopically expressed Arabidopsis. We concluded that, snRNA-seq combined with snATAC-seq is an extensible platform to explore association between the chromatin regulatory events and gene expression across diversity cell-types in peanut leaf, broadly application of this approach will enable significant advances in the functional research of tissues ontology in plant species.

Peanut pods develop underground, which is the most salient characteristic in peanut. However, its developmental transcriptome remains largely unknown. In the present study, we sequenced over one billion transcripts to explore the developmental transcriptome of peanut pod using Illumina sequencing. Moreover, we identified and quantified the abundances of 165,689 transcripts in seed and shell tissues along with a pod developmental gradient. The dynamic changes of differentially expressed transcripts (DETs) were described in seed and shell. Additionally, we found that photosynthetic genes were not only pronouncedly enriched in aerial pod, but also played roles in developing pod under dark condition. Genes functioning in photomorphogenesis showed distinct expression profiles along subterranean pod development. Clustering analysis unraveled a dynamic transcriptome, in which transcripts for DNA synthesis and cell division during pod expansion were transitioning to transcripts for cell expansion and storage activity during seed filling. Collectively, our study formed a transcriptional baseline for peanut fruit development under dark condition.

Cultivated peanut (Arachis hypogaea) is an allotetraploid crop planted in Asia, Africa, and America for edible oil and protein. To explore the origins and consequences of tetraploidy, we sequenced the allotetraploid A. hypogaea genome and compared it with the related diploid Arachis duranensis and Arachis ipaensis genomes. We annotated 39 888 A-subgenome genes and 41 526 B-subgenome genes in allotetraploid peanut. The A. hypogaea subgenomes have evolved asymmetrically, with the B subgenome resembling the ancestral state and the A subgenome undergoing more gene disruption, loss, conversion, and transposable element proliferation, and having reduced gene expression during seed development despite lacking genome-wide expression dominance. Genomic and transcriptomic analyses identified more than 2 500 oil metabolism-related genes and revealed that most of them show altered expression early in seed development while their expression ceases during desiccation, presenting a comprehensive map of peanut lipid biosynthesis. The availability of these genomic resources will facilitate a better understanding of the complex genome architecture, agronomically and economically important genes, and genetic improvement of peanut.

High oleic acid (OA) peanut seeds (PS) can be beneficial for human health. However, chemical variations in high-OA PS after domestic cooking are not fully understood. In order to investigate the impact of different cooking methods on the chemical profile of high-OA PS, widely established metabolomics approach was employed to identify the relative contents of PS metabolites. Herein, 630 metabolites within 27 categories were characterized in PS, of which 141, 157, 402 differential metabolites were observed in each treatment group (boiling, baking, and frying) when compared to the raw seed. Accordingly, bioactive substances were maximally preserved in baked high-OA PS. Further conventional methods (HPLC-UV/GC-MS) quantified the absolute composition of amino and fatty acids, verifying the reliability of metabolomic analysis. Collectively, the understanding of the phytochemical substances in relation to the domestic cooking method established a foundation for future high-OA PS processing.

High oleic acid (OA) peanut seed (PS), contains a higher ratio of oleic acid (C18:1) compared to general PS, which is favored by consumers due to its health benefits. However, comprehensive lipid metabolite profiles of high-OA PS, once they have been processed via domestic cooking methods, have never been produced. To establish a scientific guide for the selection of the most appropriate processing method for high-OA PS, lipidomics was performed to identify 706 lipid metabolites in high-OA PS following boiling, baking and frying, between the three groups, 75, 175 and 242 lipid metabolites were differentially expressed respectively. Additionally, 46 glycerolipids with C18:1 molecular were observed in the lipid profiles of the treatment groups compared to the raw sample. Further evaluation of seven lipid peroxides and six antioxidant status of each testing group suggested that boiled PS retained the highest levels of lipids and antioxidant activity. Following these findings, boiling appears to be an appropriate processing method when attempting to conserve the beneficial substances found in the PS. Finally, the levels of major free fatty acids present in high-OA PS, were jointly quantified by conventional methods (GC-MS) and lipidomic analysis. FA/C16:0 levels were similar, FA/C18:0, FA/C18:1 displayed opposite results, FA/C18:2 levels increased following frying and FA/C18:3 levels were down regulated once the PS was boiled. This indicates that GC-MS is a potential method of validation for the results of lipidomic analysis. Conclusively, this in depth understanding of lipid content in relation to domestic cooking methods has provided a foundation for the processing of high-OA peanut products.

Background Many large-effect quantitative trait loci (QTLs) for yield and disease resistance related traits have been identified in different mapping populations of peanut (Arachis hypogaea L.) under multiple environments. However, only a limited number of QTLs have been used in marker-assisted selection (MAS) because of unfavorable epistatic interactions between QTLs in different genetic backgrounds. Thus, it is essential to identify consensus QTLs across different environments and genetic backgrounds for use in MAS. Here, we used QTL meta-analysis to identify a set of consensus QTLs for yield and disease resistance related traits in peanut. Results A new integrated consensus genetic map with 5874 loci was constructed. The map comprised 20 linkage groups (LGs) and was up to a total length of 2918.62 cM with average marker density of 2.01 loci per centimorgan (cM). A total of 292 initial QTLs were projected on the new consensus map, and 40 meta-QTLs (MQTLs) for yield and disease resistance related traits were detected on four LGs. The genetic intervals of these consensus MQTLs varied from 0.20 cM to 7.4 cM, which is narrower than the genetic intervals of the initial QTLs, meaning they may be suitable for use in MAS. Importantly, a region of the map that previously co-localized multiple major QTLs for pod traits was narrowed from 3.7 cM to 0.7 cM using an overlap region of four MQTLs for yield related traits on LG A05, which corresponds to a physical region of about 630.3 kb on the A05 pseudomolecule of peanut, including 38 annotated candidate genes (54 transcripts) related to catalytic activity and metabolic process. Additionally, one major MQTL for late leaf spot (LLS) was identified in a region of about 0.38 cM. BLAST searches identified 26 candidate genes (30 different transcripts) in this region, some of which were annotated as related to regulation of disease resistance in different plant species. Conclusions Combined with the high-density marker consensus map, all the detected MQTLs could be useful in MAS. The biological functions of the 64 candidate genes should be validated to unravel the molecular mechanisms of yield and disease resistance in peanut. Electronic supplementary material The online version of this article (10.1186/s12864-018-5288-3) contains supplementary material, which is available to authorized users.

Phosphoenolpyruvate carboxylase (PEPC) is a ubiquitous cytosolic enzyme that catalyzes the irreversible β-carboxylation of phosphoenolpyruvate (PEP) in presence of HCO3− to produce oxaloacetate (OAA) during carbon fixation and photosynthesis. It is well accepted that PEPC genes are expressed in plants upon stress. PEPC also supports the biosynthesis of biocompatible osmolytes in many plant species under osmotic stress. There are five isoforms of PEPC found in peanut (Arachis hypogaea L.), namely, AhPEPC1, AhPEPC2, AhPEPC3, AhPEPC4, and AhPEPC5. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the gene expression patterns of these AhPEPC genes were different in mature seeds, stems, roots, flowers, and leaves. The expression of all the plant type PEPC (PTPCs) (AhPEPC1, AhPEPC2, AhPEPC3, and AhPEPC4) was relatively high in roots, while the bacterial type PEPC (BTPC) (AhPEPC5) showed a remarkable expression level in flowers. Principal component analysis (PCA) result showed that AhPEPC3 and AhPEPC4 are correlated with each other, indicating comparatively associations with roots, and AhPEPC5 have a very close relationship with flowers. In order to investigate the function of these AhPEPCs, the fragments of these five AhPEPC cDNA were cloned and expressed in Escherichia coli (E. coli). The recombinant proteins contained a conserved domain with a histidine site, which is important for enzyme catalysis. Results showed that protein fragments of AhPEPC1, AhPEPC2, and AhPEPC5 had remarkable expression levels in E. coli. These three recombinant strains were more sensitive at pH 9.0, and recombinant strains carrying AhPEPC2 and AhPEPC5 fragments exhibited more growth than the control strain with the presence of PEG6000. Our findings showed that the expression of the AhPEPC fragments may enhance the resistance of transformed E. coli to osmotic stress.

In order to obtain more valuable insights into the protein dynamics and accumulation of allergens in seeds during underground development, we performed a proteomic study on developing peanut seeds at seven different stages. A total of 264 proteins with altered abundance and contained at least one unique peptide was detected by matrix-assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS). All identified proteins were classified into five functional categories as level 1 and 20 secondary functional categories as level 2. Among them, 88 identified proteins (IPs) were related to carbohydrate/ amino acid/ lipid transport and metabolism, indicating that carbohydrate/amino acid/ lipid metabolism played a key role in the underground development of peanut seeds. Hierarchical cluster analysis showed that all IPs could be classified into eight cluster groups according to the abundance profiles, suggesting that the modulatory patterns of these identified proteins were complicated during seed development. The largest group contained 41 IPs, the expression of which decreased at R 2 and reached a maximum at R3 but gradually decreased from R4. A total of 14 IPs were identified as allergen-like proteins by BLAST with A genome (Arachis duranensis) or B genome (Arachis ipaensis) translated allergen sequences. Abundance profile analysis of 14 identified allergens showed that the expression of all allergen proteins was low or undetectable by 2-DE at the early stages (R1 to R4), and began to accumulate from the R5 stage and gradually increased. Network analysis showed that most of the significant proteins were involved in active metabolic pathways in early development. Real time RT-PCR analysis revealed that transcriptional regulation was approximately consistent with expression at the protein level for 8 selected identified proteins. In addition, some amino acid sequences that may be associated with new allergens were also discussed.

Aflatoxin-affected groundnut or peanut presents a major global health issue to both commercial and subsistence farming. Therefore, understanding the genetic and molecular mechanisms associated with resistance to aflatoxin production during host&ndash, pathogen interactions is crucial for breeding groundnut cultivars with minimal level of aflatoxin contamination. Here, we performed gene expression profiling to better understand the mechanisms involved in reduction and prevention of aflatoxin contamination resulting from Aspergillus flavus infection in groundnut seeds. RNA sequencing (RNA-Seq) of 16 samples from different time points during infection (24 h, 48 h, 72 h and the 7th day after inoculation) in U 4-7-5 (resistant) and JL 24 (susceptible) genotypes yielded 840.5 million raw reads with an average of 52.5 million reads per sample. A total of 1779 unique differentially expressed genes (DEGs) were identified. Furthermore, comprehensive analysis revealed several pathways, such as disease resistance, hormone biosynthetic signaling, flavonoid biosynthesis, reactive oxygen species (ROS) detoxifying, cell wall metabolism and catabolizing and seed germination. We also detected several highly upregulated transcription factors, such as ARF, DBB, MYB, NAC and C2H2 in the resistant genotype in comparison to the susceptible genotype after inoculation. Moreover, RNA-Seq analysis suggested the occurrence of coordinated control of key pathways controlling cellular physiology and metabolism upon A. flavus infection, resulting in reduced aflatoxin production.

Peanut (Arachis hypogaea L.) is a staple crop in semiarid tropical and subtropical regions. Although the genome of peanut has been fully sequenced, the current gene annotations are still incomplete...

Modern peanut contains fatty acid desaturase 2 (FAD2) mutation, which is capable of producing high oleic acid for human health. However, the dynamic changes of the lipidome regarding fad2 remain elusive in peanut seed. In the present study, 547 lipid features were identified in high- and normal-oleic peanut seeds by utilizing the mass spectrometric approach. The fad2-induced differently expressed lipids (DELs) were polarly distributed at early and maturation stages during high-oleic acid (OA) seed development. Subsequently, integration of previously published proteomic data and lipidomic data revealed that 21 proteins and 149 DELs were annotated into the triacylglycerol assembly map, of which nine enzymes and 31 lipid species shared similar variation tendencies. Additionally, the variation tendencies of 17 acyl fatty acids were described in a hypothetical biosynthetic pathway. Collectively, the understanding of the lipid composition correlated with fad2 established a foundation for future high-OA peanut breeding based on lipidomic data.

Background Microsatellites, or simple sequence repeats (SSRs), represent important DNA variations that are widely distributed across the entire plant genome and can be used to develop SSR markers, which can then be used to conduct genetic analyses and molecular breeding. Cultivated peanut (A. hypogaea L.), an important oil crop worldwide, is an allotetraploid (AABB, 2n = 4× = 40) plant species. Because of its complex genome, genomic marker development has been very challenging. However, sequencing of cultivated peanut genome allowed us to develop genomic markers and construct a high-density physical map. Results A total of 8,329,496 SSRs were identified, including 3,772,653, 4,414,961, and 141,882 SSRs that were distributed in subgenome A, B, and nine scaffolds, respectively. Based on the flanking sequences of the identified SSRs, a total of 973,984 newly developed SSR markers were developed in subgenome A (462,267), B (489,394), and nine scaffolds (22,323), with an average density of 392.45 markers per Mb. In silico PCR evaluation showed that an average of 88.32% of the SSR markers generated only one in silico-specific product in two tetraploid A. hypogaea varieties, Tifrunner and Shitouqi. A total of 39,599 common SSR markers were identified among the two A. hypogaea varieties and two progenitors, A. duranensis and A. ipaensis. Additionally, an amplification effectiveness of 44.15% was observed by real PCR validation. Moreover, a total of 1276 public SSR loci were integrated with the newly developed SSR markers. Finally, a previously known leaf spot quantitative trait locus (QTL), qLLS_T13_A05_7, was determined to be in a 1.448-Mb region on chromosome A05. In this region, a total of 819 newly developed SSR markers were located and 108 candidate genes were detected. Conclusions The availability of these newly developed and public SSR markers both provide a large number of molecular markers that could potentially be used to enhance the process of trait genetic analyses and improve molecular breeding strategies for cultivated peanut.

A first creation of high oleic acid peanut varieties by using transcription activator-like effecter nucleases (TALENs) mediated targeted mutagenesis of Fatty Acid Desaturase 2 (FAD2). Transcription activator like effector nucleases (TALENs), which allow the precise editing of DNA, have already been developed and applied for genome engineering in diverse organisms. However, they are scarcely used in higher plant study and crop improvement, especially in allopolyploid plants. In the present study, we aimed to create targeted mutagenesis by TALENs in peanut. Targeted mutations in the conserved coding sequence of Arachis hypogaea fatty acid desaturase 2 (AhFAD2) were created by TALENs. Genetic stability of AhFAD2 mutations was identified by DNA sequencing in up to 9.52 and 4.11% of the regeneration plants at two different targeted sites, respectively. Mutation frequencies among AhFAD2 mutant lines were significantly correlated to oleic acid accumulation. Genetically, stable individuals of positive mutant lines displayed a 0.5-2 fold increase in the oleic acid content compared with non-transgenic controls. This finding suggested that TALEN-mediated targeted mutagenesis could increase the oleic acid content in edible peanut oil. Furthermore, this was the first report on peanut genome editing event, and the obtained high oleic mutants could serve for peanut breeding project.

Peanut sprouts (PSs) are edible food with a high nutritional value. It is well known that different light treatments during seedling germination affect the nutritional elements of PSs. However, the comprehensive exploration of the chemical profile variation after light and dark treatments is not widely used in PSs due to the lack of high-throughout examination technology. In the present study, a widely targeted method based on UHPLC-QTRAP-MS/MS equipment was carried out to identify and screen differential metabolites. A total of 797 metabolites were identified from PSs, and 97 differential metabolites were screened between light and dark treatment groups. The majority of phytochemical compounds, such as flavonoids, alkaloids, nucleotides, amino acids, saccharides, and alcohols, were significantly up-regulated in PSs germinated under the dark conditions, which were positively related to the large-scale metabolic data of human diseases by KEGG metabolic pathway analysis. Collectively, our data suggest that the seeds should be protected from exposure to light during PS germination, which might improve the nutritional value of the final peanut product.

Peanuts with high oleic acid content are usually considered to be beneficial for human health and edible oil storage. In breeding practice, peanut lines with high monounsaturated fatty acids are selected using fatty acid desaturase 2 (FAD2), which is responsible for the conversion of oleic acid (C18:1) to linoleic acid (C18:2). Here, comparative transcriptomics were used to analyze the global gene expression profile of high- and normal-oleic peanut cultivars at six time points during seed development. First, the mutant type of FAD2 was determined in the high-oleic peanut (H176). The result suggested that early translation termination occurred simultaneously in the coding sequence of FAD2-A and FAD2-B, and the cultivar H176 is capable of utilizing a potential germplasm resource for future high-oleic peanut breeding. Furthermore, transcriptomic analysis identified 74 differentially expressed genes (DEGs) involved in lipid metabolism in high-oleic peanut seed, of which five DEGs encoded the fatty acid desaturase. Aradu.XM2MR belonged to the homologous gene of stearoyl-ACP (acyl carrier protein) desaturase 2 (SAD2) that converted the C18:0 into C18:1. Further subcellular localization studies indicated that FAD2 was located at the endoplasmic reticulum (ER), and Aradu.XM2MR was targeted to the plastid in Arabidopsis protoplast cells. To examine the dynamic mechanism of this finding, we focused on the peroxidase (POD)-mediated fatty acid (FA) degradation pathway. The fad2 mutant significantly increased the POD activity and H2O2 concentration at the early stage of seed development, implying that redox signaling likely acted as a messenger to connect the signaling transduction between the high-oleic content and Aradu.XM2MR transcription level. Taken together, transcriptome analysis revealed the feedback mechanism of SAD2 (Aradu.XM2MR) associated with FAD2 mutation during the seed developmental stage, which could provide a potential peanut breeding strategy based on identified candidate genes to improve the content of oleic acid.

Additional file 2: Figure S1. Chromosome-wide distribution of SSRs in A. hypogaea cv. Fuhuasheng genome. Figure S2. Number of SSR repeat motifs. Figure S3. Abundance of the top 30 different types of SSR motifs. Figure S4. Number (A) and percentage (B) of different types of SSR markers. Figure S5. Summary of SSR types of the developed SSR markers. Figure S6. Product size of 188 SSR markers tested by PCR amplification. Figure S7. Number of loci in the public SSR markers as determined by e-PCR remapping. Figure S8. Comparison of known QTLs in genetic and physical maps.

Peanut or groundnut (Arachis hypogaea L.), a legume of South American origin, has high seed oil content (45–56%) and is a staple crop in semiarid tropical and subtropical regions, partially because of drought tolerance conferred by its geocarpic reproductive strategy. We present a draft genome of the peanut A-genome progenitor, Arachis duranensis, and 50,324 protein-coding gene models. Patterns of gene duplication suggest the peanut lineage has been affected by at least three polyploidizations since the origin of eudicots. Resequencing of synthetic Arachis tetraploids reveals extensive gene conversion in only three seed-to-seed generations since their formation by human hands, indicating that this process begins virtually immediately following polyploid formation. Expansion of some specific gene families suggests roles in the unusual subterranean fructification of Arachis. For example, the S1Fa-like transcription factor family has 126 Arachis members, in contrast to no more than five members in other examined plant species, and is more highly expressed in roots and etiolated seedlings than green leaves. The A. duranensis genome provides a major source of candidate genes for fructification, oil biosynthesis, and allergens, expanding knowledge of understudied areas of plant biology and human health impacts of plants, informing peanut genetic improvement and aiding deeper sequencing of Arachis diversity.

A characteristic feature of peanut is the subterranean fructification, geocarpy, in which the gynophore ('peg'), a specialized organ that transitions from upward growth habit to downward outgrowth upon fertilization, drives the developing pod into the soil for subsequent development underground. As a step towards understanding this phenomenon, we explore the developmental dynamics of the peanut pod transcriptome at 11 successive stages. We identified 110 217 transcripts across developmental stages and quantified their abundance along a pod developmental gradient in pod wall. We found that the majority of transcripts were differentially expressed along the developmental gradient as well as identified temporal programs of gene expression, including hundreds of transcription factors. Thought to be an adaptation to particularly harsh subterranean environments, both up- and down-regulated gene sets in pod wall were enriched for response to a broad array of stimuli, like gravity, light and subterranean environmental factors. We also identified hundreds of transcripts associated with gravitropism and photomorphogenesis, which may be involved in the geocarpy. Collectively, this study forms a transcriptional baseline for geocarpy in peanut as well as provides a considerable body of evidence that transcriptional regulation in peanut aerial and subterranean fruits is complex.

In the original article, there was an error. The link of sequencing data was not attached in Supplementary Material section. A correction has been made to the Supplementary Material section: The genome assembly and annotation data were deposited in the DRYAD digital repository (https://datadryad.org/). All the data can be downloaded from DRYAD by searching for doi: 10.5061/dryad.hm5vs13 or by directly accessing the link: https://doi.org/10.5061/dryad.hm5vs13. The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way. The original article has been updated.

Peanuts (Arachis hypogaea L.) are an important oilseed crop, containing high contents of protein and fatty acids (FA). The major components of FA found in peanut oil are unsaturated FAs, including oleic acid (OA, C18:1) and linoleic acid (LOA, C18:2). Moreover, the high content of OA in peanut oil is beneficial for human health and long-term storage due to its antioxidant activity. However, the dynamic changes in proteomics related to OA accumulation during seed development still remain largely unexplored. In the present study, a comparative proteome analysis based on iTRAQ (isobaric Tags for Relative and Absolute Quantification) was performed to identify the critical candidate factors involved in OA formation. A total of 389 differentially expressed proteins (DEPs) were identified between high-oleate cultivar Kainong176 and low-oleate cultivar Kainong70. Among these DEPs, 201 and 188 proteins were upregulated and downregulated, respectively. In addition, these DEPs were categorized into biosynthesis pathways of unsaturated FAs at the early stage during the high-oleic peanut seed development, and several DEPs involved in lipid oxidation pathway were found at the stage of seed maturation. Meanwhile, 28 DEPs were sporadically distributed in distinct stages of seed formation, and their molecular functions were directly correlated to FA biosynthesis and degradation. Fortunately, the expression of FAB2 (stearoyl-acyl carrier protein desaturase), the rate-limiting enzyme in the upstream biosynthesis process of OA, was significantly increased in the early stage and then decreased in the late stage of seed development in the high-oleate cultivar Kainong176. Furthermore, real-time PCR verified the expression pattern of FAB2 at the mRNA level, which was consistent with its protein abundance. However, opposite results were found for the low-oleate cultivar Kainong70. Overall, the comparative proteome analysis provided valuable insight into the molecular dynamics of OA accumulation during peanut seed development.

Peanuts (Arachis hypogaea L.) are an important oilseed crop, containing high contents of protein and fatty acids (FA). The major components of FA found in peanut oil are unsaturated FAs, including oleic acid (OA, C18:1) and linoleic acid (LOA, C18:2). Moreover, the high content of OA in peanut oil is beneficial for human health and long-term storage due to its antioxidant activity. However, the dynamic changes in proteomics related to OA accumulation during seed development still remain largely unexplored. In the present study, a comparative proteome analysis based on iTRAQ (isobaric Tags for Relative and Absolute Quantification) was performed to identify the critical candidate factors involved in OA formation. A total of 389 differentially expressed proteins (DEPs) were identified between high-oleate cultivar Kainong176 and low-oleate cultivar Kainong70. Among these DEPs, 201 and 188 proteins were upregulated and downregulated, respectively. In addition, these DEPs were categorized into biosynthesis pathways of unsaturated FAs at the early stage during the high-oleic peanut seed development, and several DEPs involved in lipid oxidation pathway were found at the stage of seed maturation. Meanwhile, 28 DEPs were sporadically distributed in distinct stages of seed formation, and their molecular functions were directly correlated to FA biosynthesis and degradation. Fortunately, the expression of FAB2 (stearoyl-acyl carrier protein desaturase), the rate-limiting enzyme in the upstream biosynthesis process of OA, was significantly increased in the early stage and then decreased in the late stage of seed development in the high-oleate cultivar Kainong176. Furthermore, real-time PCR verified the expression pattern of FAB2 at the mRNA level, which was consistent with its protein abundance. However, opposite results were found for the low-oleate cultivar Kainong70. Overall, the comparative proteome analysis provided valuable insight into the molecular dynamics of OA accumulation during peanut seed development.

Peanut (Arachis hypogaea L.), an important leguminous crop, is widely cultivated in tropical and subtropical regions. Peanut is an allotetraploid, having A and B subgenomes that maybe have originated in its diploid progenitors Arachis duranensis (A-genome) and Arachis ipaensis (B-genome), respectively. We previously sequenced the former and here present the draft genome of the latter, expanding our knowledge of the unique biology of Arachis. The assembled genome of A. ipaensis is ~1.39 Gb with 39,704 predicted protein-encoding genes. A gene family analysis revealed that the FAR1 family may be involved in regulating peanut special fruit development. Genomic evolutionary analyses estimated that the two progenitors diverged ~3.3 million years ago and suggested that A. ipaensis experienced a whole-genome duplication event after the divergence of Glycine max. We identified a set of disease resistance-related genes and candidate genes for biological nitrogen fixation. In particular, two and four homologous genes that may be involved in the regulation of nodule development were obtained from A. ipaensis and A. duranensis, respectively. We outline a comprehensive network involved in drought adaptation. Additionally, we analyzed the metabolic pathways involved in oil biosynthesis and found genes related to fatty acid and triacylglycerol synthesis. Importantly, three new FAD2 homologous genes were identified from A. ipaensis and one was completely homologous at the amino acid level with FAD2 from A. hypogaea. The availability of the A. ipaensis and A. duranensis genomic assemblies will advance our knowledge of the peanut genome.

Figure S3. Enrichment analysis of gene ontology terms for the candidate genes for yield and late leaf spot. (DOCX 174â kb)

Figure S1. Distribution of all the initial QTLs on the linkage groups of the integrated consensus map. (DOCX 671â kb)

Peanut (Arachis hypogaea L.) is one of the most important oil-bearing crops in the world. The gravitropic response of peanut gynophores plays an essential role in peanut reproductive development. In this study, we developed an in vitro culture system and applied it to the study of peanut gynophore gravitropism. By comparing the proteomes of gynophores grown in vitro with the tip pointing upward (gravity stimulation sample) and downward (natural growth control) at 6 h and 12 h, we observed 42 and 39 with significantly altered expression pattern at 6 and 12 h, respectively. Out of these proteins, 13 proteins showed same expression profiling at both 6 h and 12 h. They were identified by MALDI-TOF/TOF and further characterized with quantitative real time RT-PCR. Among the 13 identified proteins, two were identified as class III acidic endochitinases, two were identified as Kunitz trypsin protease inhibitors, and the remaining proteins were identified as pathogenesis-related class 10 protein, Ara h 8 allergen isoform 3, voltage-dependent anion channel, gamma carbonic anhydrase 1, germin-like protein subfamily 3 member 3 precursor, chloride channel, glycine-rich RNA-binding protein and gibberellin receptor GID1. Real time RT-PCR analysis revealed that transcriptional regulation is consistent with expression at the protein level for class III acidic endochitinase, Kunitz trypsin protease inhibitor, chloride channel and pathogenesis-related class 10 protein, while the expression of the other 7 proteins might be regulated at post-transcriptional levels. This study identified several potential gravitropic response proteins in peanut gynophores and helps to understand early gravitropic responses in peanut gynophores. Biological significance The gravitropic response of the peanut gynophores plays an essential role in peanut production. However, the molecular mechanism responsible for gravitropic responses in the peanut gynophores has not been explored yet. The result generated in this study may provide in vitro culture system for gravitropism study of plant gravitropic response and novel insights into the proteome-level response and give a more comprehensive understanding of early gravitropic response in peanut gynophores. This article is part of a Special Issue entitled: Translational Plant Proteomics.

The peanut plant produces flowers aerially, while develops the fruits and seeds underground. Pod swelling is a vital process of peanut pod and seed development only occurring after the gynophore carrying the ovule into the soil. The failure of gynophore penetration into the soil leads to suppression of pod swelling initiation. However, the molecular mechanism underlying the process remains unknown. A comparative proteome analysis between developing aerial and subterranean pods at various developmental stages was performed using 2-DE approach. 47 significantly differentially expressed spots were selected to further identification by MALDI-TOF–TOF MS. They were corresponded to 31 distinct proteins, suggesting that many identified spots were modified in post-translation. Functional annotation revealed their involvement in twelve important biological processes, such as photosynthesis, oxidative stress response, lignin synthesis, fatty acid biosynthesis, glycolysis, protein catabolic process, cellular metabolic process, regulation process, etc. Furthermore, 10 identified proteins were validated by real-time RT-PCR analysis. Several photosynthesis and oxidative stress proteins displayed elevated expression levels in aerial pods. Otherwise, enzymes in lignin synthesis and ubiquitin proteasome system were down-accumulation in subterranean pods. These enzymes might function as potential candidate proteins and play critical roles to regulate pods swelling and development. Biological significance Pod swelling plays a crucial role in peanut fruit and seed development. However, a large number of aerial pods can't form normal pods due to suppression of swelling initiation by the failure of penetration into the soil, thereby causing to seed yield loss. Limited knowledge is available underlying molecular mechanism regulating initiation of swelling in peg tips and pod development. The results generated in this study may provide evidence for some functional proteins as potential candidates to pod swelling and new molecular insights to improve our understanding of pod development under light and darkness conditions, which may contribute valuable information to high yield breeding in future.