Cultivated Peanut Research Articles

Evaluation and Validation of Reference Genes for Normalization of Quantitative Real-Time PCR Based Gene Expression Studies in Peanut

The quantitative real-time PCR (qPCR) based techniques have become essential for gene expression studies and high-throughput molecular characterization of transgenic events. Normalizing to reference gene in relative quantification make results from qPCR more reliable when compared to absolute quantification, but requires robust reference genes. Since, ideal reference gene should be species specific, no single internal control gene is universal for use as a reference gene across various plant developmental stages and diverse growth conditions. Here, we present validation studies of multiple stably expressed reference genes in cultivated peanut with minimal variations in temporal and spatial expression when subjected to various biotic and abiotic stresses. Stability in the expression of eight candidate reference genes including ADH3, ACT11, ATPsyn, CYP2, ELF1B, G6PD, LEC and UBC1 was compared in diverse peanut plant samples. The samples were categorized into distinct experimental sets to check the suitability of candidate genes for accurate and reliable normalization of gene expression using qPCR. Stability in expression of the references genes in eight sets of samples was determined by geNorm and NormFinder methods. While three candidate reference genes including ADH3, G6PD and ELF1B were identified to be stably expressed across experiments, LEC was observed to be the least stable, and hence must be avoided for gene expression studies in peanut. Inclusion of the former two genes gave sufficiently reliable results; nonetheless, the addition of the third reference gene ELF1B may be potentially better in a diverse set of tissue samples of peanut.

Genetic Diversity of <I>Arachis</I> Accessions Detected by EST-SSR from Cultivated Peanut (<I>Arachis hypogaea</I> L.)

The amplification efficiency and application potential for the research on peanut genetic diversity by using cultivars-derived EST-SSR primers in 89 Arachis aeecssions were assessed.Based on randomly selected 235 EST-SSR primer pairs,we detected 223 amplification bands in the 89 accessions with the amplification efficiency of 94.89%.Among them,53 primer pairs exhibited 206 polymorphic bands,which showed 1-12 bands per primer pair with an average of 3.89.The polymorphism index ranged from 0.044 to 4.040 with an average of 1.173.The average similarity coefficient was 0.685 among 89 peanut accessions,ranging from 0.442 to 0.976.The accessions from the same section were mainly clustered together,peanut cultivars were grouped into the section of Arachis,and A.duranensis had a close relation to cultivated peanut,which is coincident with botanical classification.According to the statistical analysis from the results of oil content and SSR data in wild peanuts,some interesting bands such as POCR437-180/170 were found to be potential for candidate markers related to oil content.BLAST analysis indicated that the alignment sequence of primer POCR437 corresponded to the coding genes of malonyl-CoA-ACP transacylase and acyl-carrier-protein,both of which are involved in fatty acid synthesis.

An AS-PCR assay for accurate genotyping of <i>FAD2A/FAD2B</i> genes in peanuts (<i>Arachis hypogaea</i> L.)

FAD2A and FAD2B are homoeologous genes from A and B genomes in cultivated peanuts (Arachis hypogaea L.) encoding fatty acid desaturates which convert oleate to linoleate. To study the genetics of oleate and breed high oleate peanut cultivars, a simple allele specific-PCR (AS-PCR) protocol for the accurate genotyping of FAD2A/FAD2B was developed to discriminate the wild and mutant allele of both genes (FAD2A 448 G > A and FAD2B 441_442insA). The results may serve to develop a feasible procedure for producing highly desired high oleate peanut cultivars through hybridization.

Recent Advances in Molecular Genetic Linkage Maps of Cultivated Peanut

ABSTRACTThe competitiveness of peanuts in domestic and global markets has been threatened by losses in productivity and quality that are attributed to diseases, pests, environmental stresses and allergy or food safety issues. Narrow genetic diversity and a deficiency of polymorphic DNA markers severely hindered construction of dense genetic maps and quantitative trait loci (QTL) mapping in order to deploy linked markers in marker-assisted peanut improvement. The U.S. Peanut Genome Initiative (PGI) was launched in 2004, and expanded to a global effort in 2006 to address these issues through coordination of international efforts in genome research beginning with molecular marker development and improvement of map resolution and coverage. Ultimately, a peanut genome sequencing project was launched in 2012 by the Peanut Genome Consortium (PGC). We reviewed the progress for accelerated development of peanut genomic resources in peanut, such as generation of expressed sequenced tags (ESTs) (252,832 ESTs as December 2012 in the public NCBI EST database), development of molecular markers (over 15,518 SSRs), and construction of peanut genetic linkage maps, in particular for cultivated peanut. Several consensus genetic maps have been constructed, and there are examples of recent international efforts to develop high density maps. An international reference consensus genetic map was developed recently with 897 marker loci based on 11 published mapping populations. Furthermore, a high-density integrated consensus map of cultivated peanut and wild diploid relatives also has been developed, which was enriched further with 3693 marker loci on a single map by adding information from five new genetic mapping populations to the published reference consensus map.

Characterization of resveratrol content in ten wild species of section Arachis, genus Arachis

Peanut is one of the few plants that synthesizes resveratrol, a phenolic compound of the stilbene class, which has been associated with reduced risk of developing chronic diseases, such as cancer, cardiovascular diseases, skin diseases, pulmonary diseases, diabetes and neurological diseases. Resveratrol was detected in different parts of the peanut plant, including roots, leaves, seeds and their derivatives. The wild species of the Arachis section are also strong candidates to synthesize resveratrol because they are phylogenetically closely related to cultivated peanut. Our objective was to characterize the resveratrol content in ten wild species of Arachis with three different genomes (A, B and K). The plant material was composed of leaves of the ten species treated (test) and not treated (control) with ultraviolet (UV) radiation. The test and control samples were extracted and the identification and quantification of resveratrol was performed using high performance liquid chromatography (HPLC). All species studied synthesized resveratrol and the concentrations ranged from 299.5 μg/g in A. kempff-mercadoi to 819.9 μg/g in A. cardenasii. DPPH antioxidant activity varied between 18.7 % for A. duranensis and 48.2 % in A. simpsonii. The results showed that wild Arachis species are a potential source of alleles for improvement of cultivated peanut, with the aim of achieving higher resveratrol content in leaves.

Generation Means Analysis of Oil Concentration in Peanut

The current interest in biodiesel production has resulted in a concurrent interest in increasing the oil concentration in high-yielding cultivars, which could make peanuts (Arachis hypogaea.) more desirable as a biofuel source. Currently, peanut seed is approximately 450 to 500 g kg−1 oil on a dry weight basis, depending upon location grown, and there is relatively little genetic variation for oil concentration among adapted high-yielding cultivated peanut genotypes. Thus, identifying sources of variation and elucidating the genetics of oil concentration in peanut is essential to advancing the development of high oil genotypes. The objective of this study was to determine the types of gene action governing the inheritance of oil concentration in peanut by generation means analysis. The F1, F2, and backcross generations of two different runner peanut crosses segregating for oil concentration were evaluated in College Station, Texas, in 2010. Significant differences in oil concentration among the generations were detected, and generation means analysis revealed significant additive, dominance, and epistatic effects for oil concentration in both crosses. The broad-sense heritability estimates were 0.85 and 0.78, and narrow-sense heritability estimates were 0.55 and 0.53 for each of the crosses. Our data indicate that transgressive segregants for high oil were observed, and there is sufficient additive variation present to improve the oil concentration of current runner cultivars.

Biochemical Composition and Disease Resistance in Newly Synthesized Amphidiploid and Autotetraploid Peanuts

Genetic diversity in peanut (Arachishypogaea L.) is narrow due to its evolution and domestication processes. Amphidiploids and autotetraploids (newly synthesized tetraploids) were created to broaden its genetic base. Molecular analysis has shown that the newly synthesized tetraploids had broader genetic base; and were genetically divergent when compared to cultivated peanut. Nutritional composition relative to oil, fatty acid composition, O/L ratio, protein, iodine value and presence of plant proteinase inhibitors such as trypsin and chymotrypsin inhibitors were studied in the synthesized tetraploids. Some of the newly synthesized tetraploids had higher amounts of proteinase inhibitors. Evaluation of newly synthesized tetraploids revealed several lines resistant to late leaf spot (LLS) and peanut bud necrosis disease (PBND).

An electrospray ionisation-mass spectrometry screening of triacylglycerols in developing cultivated and wild peanut kernels (Arachis hypogaea L.)

The accumulation of triacylglycerols during the development of three varieties of peanuts was monitored in two Tunisian cultivated peanut (Trabelsia (AraT) and Chounfakhi (AraC)) and one wild Tunisian peanut (Arbi (AraA)). The presence of TAGs composed of rare fatty acid residues such as hexacosanoic acid (C23:0) and heneicosanoic acid (C21:0) among the triacylglycerols C23:0 LL, C23:0 OO and C21:0 LL was noted. The major molecular species of triacylglycerol detected in the three peanut varieties were dioleoyl linoleoyl (OOL), 1,2,3-trioleyl (OOO), 1,2-dioleyl-3-palmitoyl (POO), 1,2-dilinoleoyl-3-oleyl (OLL) and 1-oleoyl-2-linoleoyl-3-linolenoyl (OLLn). The TAG composition and content were significantly different among the three peanut varieties. The three major TAGs were OOL (20.6%), OOO (15.6%) and OLLn (13.2%) in AraA; OOL (21.4%), OOO (20.1%) and POO (17.5%) in AraC and finally OLL (20.7%), OOO (19.8%) and OLL (17.7%) in AraT.

A study of the relationships of cultivated peanut (Arachis hypogaea) and its most closely related wild species using intron sequences and microsatellite markers

The genus Arachis contains 80 described species. Section Arachis is of particular interest because it includes cultivated peanut, an allotetraploid, and closely related wild species, most of which are diploids. This study aimed to analyse the genetic relationships of multiple accessions of section Arachis species using two complementary methods. Microsatellites allowed the analysis of inter- and intraspecific variability. Intron sequences from single-copy genes allowed phylogenetic analysis including the separation of the allotetraploid genome components. Intron sequences and microsatellite markers were used to reconstruct phylogenetic relationships in section Arachis through maximum parsimony and genetic distance analyses. Although high intraspecific variability was evident, there was good support for most species. However, some problems were revealed, notably a probable polyphyletic origin for A. kuhlmannii. The validity of the genome groups was well supported. The F, K and D genomes grouped close to the A genome group. The 2n = 18 species grouped closer to the B genome group. The phylogenetic tree based on the intron data strongly indicated that A. duranensis and A. ipaënsis are the ancestors of A. hypogaea and A. monticola. Intron nucleotide substitutions allowed the ages of divergences of the main genome groups to be estimated at a relatively recent 2·3-2·9 million years ago. This age and the number of species described indicate a much higher speciation rate for section Arachis than for legumes in general. The analyses revealed relationships between the species and genome groups and showed a generally high level of intraspecific genetic diversity. The improved knowledge of species relationships should facilitate the utilization of wild species for peanut improvement. The estimates of speciation rates in section Arachis are high, but not unprecedented. We suggest these high rates may be linked to the peculiar reproductive biology of Arachis.

Molecular cloning and characterization of an acyl-ACP thioesterase gene (AhFatB1) from allotetraploid peanut (Arachis hypogaea L.)

Acyl-acyl carrier protein (ACP) thioesterase is a nuclear encoded plastid localized enzyme which plays an essential role in chain termination during de novo fatty acid synthesis in plant. FatB genes coding for this enzyme from a variety of plant species have been isolated and characterized. However, there are few reports on such genes in peanut ( Arachis hypogaea ), an important edible and oilseed crop. In this study, full-length cDNA of an acyl-acyl carrier protein thioesterase (EC 3.1.2.14), designated as AhFatB1 , was isolated from peanut cDNA libraries. The putative open reading frames consist of 1239 bp with five introns spliced from the corresponding genomic sequence, encoding a 413 amino acid protein, two homologous genes, AhFatB1A and AhFatB1B , with sequence difference at the 5’ non-coding regions were characterized at the nucleotide level from different cultivated peanut genotypes, and the two genes have their origin in different diploid progenitor which was evidenced by the characterization of AhFatB1 genes from Arachis duranensis and Arachis ipaensis , the putative A-genome donor and B-genome donor respectively. AhFatB1 genes are constitutively expressed in peanut tissues and the total FatB1 transcript accumulations are temporally regulated during peanut seed development. Keywords: Peanut thioesterase, palmitic acid oilseed

Comparison of gene expression profiles in cultivated peanut (Arachis hypogaea) under strong artificial selection

With 9 figures and 1 tableAbstractOver the past five decades, cultivated peanut in China has been subjected to strong artificial selection in breeding programmes. To investigate the impact of artificial selection on expression diversity, we compared gene expression profiles in pod and leaf of five widespread cultivars in Southern China. In terms of tissues, hierarchical clustering analysis revealed that expression data of pod and leaf generated different dendrograms owing to artificial selection. K‐means analysis also showed that there were 16 gene expression patterns in leaf, while only eight in pod. In considering cultivars, a cultivar specificity index (τ) was employed to characterize expression patterns, which suggested that genes having 0.15 < τ < 0.85 constituted >80% of all expression patterns. Additionally, the diversity of gene expression in pod among cultivars of the ‘YY7’ pedigree decreased from 23.8% to 3.9%. Taken together, nucleotide polymorphisms in regulatory elements owing to artificial selection led to low‐expression polymorphisms in both tissues and cultivars, contributed to the narrow genetic diversity and might be a driving force behind the breeding of cultivated peanut.

Cloning and Characterization of Chromosomal Markers from a Cot-1 Library of Peanut (Arachis hypogaea L.)

The cultivated peanut, Arachis hypogaea (AABB, 2n = 40), is an allotetraploid which was probably originated from a hybridization event between 2 ancestors, A. duranensis (A genome) and A. ipaensis (B genome) followed by chromosome doubling. The wild species in the Arachis section are useful genetic resources for genes that confer biotic and abiotic stress resistance for peanut breeding. However, the resource is not well exploited because little information on the genetic, cytogenetic, and phylogenetic relationships between cultivated peanut and its wild relatives is known. Characterization of its chromosome components will benefit the understanding of these issues. But the paucity of information on the DNA sequence and the presence of morphologically similar chromosomes impede the construction of a detailed karyotype for peanut chromosome identification. In our study, a peanut Cot-1 library was constructed to isolate highly and moderately repetitive sequences from the cultivated peanut, and the chromosomal distributions of these repeats were investigated. Both genome and chromosome specific markers were identified that allowed the distinguishing of A and B genomes in tetraploid peanut and a possible karyotyping of peanut chromosomes by FISH. In particular, a 115-bp tandem repetitive sequence was identified to be a possible centromere repetitive DNA, mainly localized in the centromeres of B chromosomes, and a partial retrotransposable element was also identified in the centromeres of B chromosomes. The cloning and characterization of various chromosomal markers is a major step for FISH-based karyotyping of peanut. The FISH markers are expected to provide a reference tool for sequence assembly, phylogenetic studies of peanut and its wild species, and breeding.

Isolation and characterization of a stress responsive small GTP-binding protein AhRabG3b in peanut (Arachis hypogaea L.)

Rab7 is a member of small GTP-binding protein family involved in intracellular vesicle trafficking from late endosome to vacuole, some of them are found to be responsive by different environmental stresses. Here we report the isolation of a Rab7 (RabG3b) cDNA from peanut (Arachis hypogaea). Overexpression of the open reading frame (ORF) of Rab7 cDNA encoding 205 amino acids in E. coli conferred the cells tolerance to various abiotic stresses. Transcription of AhRabG3b was differentially up-regulated by different environmental stimuli such as cold, dehydration, NaCl and the plant hormone ABA. Overexpression of the AhRabG3b exhibited enhanced tolerance to drought and salinity stresses in transgenic peanut. These results suggested that the AhRabG3b be a potential candidate gene responsible for both salinity and dehydration tolerance in cultivated peanut.

Expressed sequence tags in cultivated peanut (Arachis hypogaea): discovery of genes in seed development and response to Ralstonia solanacearum challenge

Although an important oil crop, peanut has only 162,030 expressed sequence tags (ESTs) publicly available, 86,943 of which are from cultivated plants. More ESTs from cultivated peanuts are needed for isolation of stress-resistant, tissue-specific and developmentally important genes. Here, we generated 63,234 ESTs from our 5 constructed peanut cDNA libraries of Ralstonia solanacearum challenged roots, R. solanacearum challenged leaves, and unchallenged cultured peanut roots, leaves and developing seeds. Among these ESTs, there were 14,547 unique sequences with 7,961 tentative consensus sequences and 6,586 singletons. Putative functions for 47.8% of the sequences were identified, including transcription factors, tissue-specific genes, genes involved in fatty acid biosynthesis and oil formation regulation, and resistance gene analogue genes. Additionally, differentially expressed genes, including those involved in ethylene and jasmonic acid signal transduction pathways, from both peanut leaves and roots, were identified in R. solanacearum challenged samples. This large expression dataset from different peanut tissues will be a valuable source for marker development and gene expression analysis. It will also be helpful for finding candidate genes for fatty acid synthesis and oil formation regulation as well as for studying mechanisms of interactions between the peanut host and R. solanacearum pathogen.

The effect of tetraploidization of wild Arachis on leaf morphology and other drought-related traits

Cultivated peanut is an allotetraploid (genome type AABB) with a very narrow genetic base, therefore wild species are an attractive source of new variability and traits. Because most wild species are diploid, the first step of introgression usually involves hybridization of wild species and polyploidization to produce a synthetic allotetraploid (AABB) that is sexually compatible with peanut. This study investigates drought-related traits such as leaf morphology, transpiration profile, chlorophyll meter readings (SCMR), specific leaf area (SLA) and transpiration rate per leaf area for two wild diploids (Arachis duranensis and Arachis ipaënsis) that could be of interest for improvement of the peanut crop. Furthermore, the inheritance of the traits from the diploid to the tetraploid state was investigated. Results showed that whilst some diploid traits such as SCMR, are maintained through hybridization and polyploidization, most characters, such as the leaf area, stomata size, trichome density and transpiration profile, are substantially modified. The study concludes that direct evaluations of drought-related traits in wild diploids may be useful for evaluation of wild species to be used in introgression. However, evaluations on wild-derived synthetic tetraploids are likely to be more informative.

Fostered and left behind alleles in peanut: interspecific QTL mapping reveals footprints of domestication and useful natural variation for breeding.

BackgroundPolyploidy can result in genetic bottlenecks, especially for species of monophyletic origin. Cultivated peanut is an allotetraploid harbouring limited genetic diversity, likely resulting from the combined effects of its single origin and domestication. Peanut wild relatives represent an important source of novel alleles that could be used to broaden the genetic basis of the cultigen. Using an advanced backcross population developed with a synthetic amphidiploid as donor of wild alleles, under two water regimes, we conducted a detailed QTL study for several traits involved in peanut productivity and adaptation as well as domestication.ResultsA total of 95 QTLs were mapped in the two water treatments. About half of the QTL positive effects were associated with alleles of the wild parent and several QTLs involved in yield components were specific to the water-limited treatment. QTLs detected for the same trait mapped to non-homeologous genomic regions, suggesting differential control in subgenomes as a consequence of polyploidization. The noteworthy clustering of QTLs for traits involved in seed and pod size and in plant and pod morphology suggests, as in many crops, that a small number of loci have contributed to peanut domestication.ConclusionIn our study, we have identified QTLs that differentiated cultivated peanut from its wild relatives as well as wild alleles that contributed positive variation to several traits involved in peanut productivity and adaptation. These findings offer novel opportunities for peanut improvement using wild relatives.

Development and characterization of highly polymorphic long TC repeat microsatellite markers for genetic analysis of peanut

BackgroundPeanut (Arachis hypogaea L.) is a crop of economic and social importance, mainly in tropical areas, and developing countries. Its molecular breeding has been hindered by a shortage of polymorphic genetic markers due to a very narrow genetic base. Microsatellites (SSRs) are markers of choice in peanut because they are co-dominant, highly transferrable between species and easily applicable in the allotetraploid genome. In spite of substantial effort over the last few years by a number of research groups, the number of SSRs that are polymorphic for A. hypogaea is still limiting for routine application, creating the demand for the discovery of more markers polymorphic within cultivated germplasm.FindingsA plasmid genomic library enriched for TC/AG repeats was constructed and 1401 clones sequenced. From the sequences obtained 146 primer pairs flanking mostly TC microsatellites were developed. The average number of repeat motifs amplified was 23. These 146 markers were characterized on 22 genotypes of cultivated peanut. In total 78 of the markers were polymorphic within cultivated germplasm. Most of those 78 markers were highly informative with an average of 5.4 alleles per locus being amplified. Average gene diversity index (GD) was 0.6, and 66 markers showed a GD of more than 0.5. Genetic relationship analysis was performed and corroborated the current taxonomical classification of A. hypogaea subspecies and varieties.ConclusionsThe microsatellite markers described here are a useful resource for genetics and genomics in Arachis. In particular, the 66 markers that are highly polymorphic in cultivated peanut are a significant step towards routine genetic mapping and marker-assisted selection for the crop.

Development and characterization of BAC-end sequence derived SSRs, and their incorporation into a new higher density genetic map for cultivated peanut (Arachis hypogaea L.).

BackgroundCultivated peanut (Arachis hypogaea L.) is an important crop worldwide, valued for its edible oil and digestible protein. It has a very narrow genetic base that may well derive from a relatively recent single polyploidization event. Accordingly molecular markers have low levels of polymorphism and the number of polymorphic molecular markers available for cultivated peanut is still limiting.ResultsHere, we report a large set of BAC-end sequences (BES), use them for developing SSR (BES-SSR) markers, and apply them in genetic linkage mapping. The majority of BESs had no detectable homology to known genes (49.5%) followed by sequences with similarity to known genes (44.3%), and miscellaneous sequences (6.2%) such as transposable element, retroelement, and organelle sequences. A total of 1,424 SSRs were identified from 36,435 BESs. Among these identified SSRs, dinucleotide (47.4%) and trinucleotide (37.1%) SSRs were predominant. The new set of 1,152 SSRs as well as about 4,000 published or unpublished SSRs were screened against two parents of a mapping population, generating 385 polymorphic loci. A genetic linkage map was constructed, consisting of 318 loci onto 21 linkage groups and covering a total of 1,674.4 cM, with an average distance of 5.3 cM between adjacent loci. Two markers related to resistance gene homologs (RGH) were mapped to two different groups, thus anchoring 1 RGH-BAC contig and 1 singleton.ConclusionsThe SSRs mined from BESs will be of use in further molecular analysis of the peanut genome, providing a novel set of markers, genetically anchoring BAC clones, and incorporating gene sequences into a linkage map. This will aid in the identification of markers linked to genes of interest and map-based cloning.

In silico polymorphism analysis for the development of simple sequence repeat and transposon markers and construction of linkage map in cultivated peanut

BackgroundPeanut (Arachis hypogaea) is an autogamous allotetraploid legume (2n = 4x = 40) that is widely cultivated as a food and oil crop. More than 6,000 DNA markers have been developed in Arachis spp., but high-density linkage maps useful for genetics, genomics, and breeding have not been constructed due to extremely low genetic diversity. Polymorphic marker loci are useful for the construction of such high-density linkage maps. The present study used in silico analysis to develop simple sequence repeat-based and transposon-based markers.ResultsThe use of in silico analysis increased the efficiency of polymorphic marker development by more than 3-fold. In total, 926 (34.2%) of 2,702 markers showed polymorphisms between parental lines of the mapping population. Linkage analysis of the 926 markers along with 253 polymorphic markers selected from 4,449 published markers generated 21 linkage groups covering 2,166.4 cM with 1,114 loci. Based on the map thus produced, 23 quantitative trait loci (QTLs) for 15 agronomical traits were detected. Another linkage map with 326 loci was also constructed and revealed a relationship between the genotypes of the FAD2 genes and the ratio of oleic/linoleic acid in peanut seed.ConclusionsIn silico analysis of polymorphisms increased the efficiency of polymorphic marker development, and contributed to the construction of high-density linkage maps in cultivated peanut. The resultant maps were applicable to QTL analysis. Marker subsets and linkage maps developed in this study should be useful for genetics, genomics, and breeding in Arachis. The data are available at the Kazusa DNA Marker Database (http://marker.kazusa.or.jp).

Matita, a new retroelement from peanut: characterization and evolutionary context in the light of the Arachis A–B genome divergence

Cultivated peanut is an allotetraploid with an AB-genome. In order to learn more of the genomic structure of peanut, we characterized and studied the evolution of a retrotransposon originally isolated from a resistance gene analog (RGA)-containing bacterial artificial chromosome (BAC) clone. It is a moderate copy number Ty1-copia retrotransposon from the Bianca lineage and we named it Matita. Fluorescent in situ hybridization (FISH) experiments showed that Matita is mainly located on the distal regions of chromosome arms and is of approximately equal frequency on both A- and B-chromosomes. Its chromosome-specific hybridization pattern facilitates the identification of individual chromosomes, a useful cytogenetic tool considering that chromosomes in peanut are mostly metacentric and of similar size. Phylogenetic analysis of Matita elements, molecular dating of transposition events, and an estimation of the evolutionary divergence of the most probable A- and B-donor species suggest that Matita underwent its last major burst of transposition activity at around the same time of the A- and B-genome divergence about 3.5million years ago. By probing BAC libraries with overgos probes for Matita, resistance gene analogues, and single- or low-copy genes, it was demonstrated that Matita is not randomly distributed in the genome but exhibits a significant tendency of being more abundant near resistance gene homologues than near single-copy genes. The described work is a further step towards broadening the knowledge on genomic and chromosomal structure of peanut and on its evolution.