Cultivated Peanut Research Articles

Development and Characterization of EST-SSR Markers from NCBI and cDNA Library in Cultivated Peanut (<i>Arachis hypogaea L.</i>)

86 132 ESTs downloaded from GenBank in NCBI and 12 501 ESTs from cDNA library constructed by high-oil linoleic acid accession E12 were analysed. After the preprocession, there were 18 051 singletons and 9 972 contigs in the GenBank of NCBI and cDNA library. Totally 3 104 SSR locis had been screened by MISA software, accounting for 11.08% for these non-redundant ESTs. All SSR locis are divided into di-nucleotide, thi-nucleotide, tetra-nucleotide, penta-nucleotide, hexa-nucleotide and multi-nucleotide etc., and thi-nucleotide motif is the most motifs and the frequency was 43.0% and 56.8% in NCBI and cDNA libraray, respectively. The number of di- and penta-nucleotide motifs were second and third in all motifs. And the hexa-nucleotide was the least motif both in NCBI and cDNA library. In all repeat motifs nucleotide, AG/TC was the most motifs and accounted for 8.65% and 13.42% in NCBI and cDNA library respectively. Among the tri-nucleotide repeats, CTT/GAA was the most frequent motif, accounting for 6.7% and 13.42%, respectively. The repeat unit number of SSR locis is between 4 and 51.

Phylogenetic analyses of peanut resistance gene candidates and screening of different genotypes for polymorphic markers

The nucleotide-binding-site-leucine-rich-repeat (NBS–LRR)-encoding gene family has attracted much research interest because approximately 75% of the plant disease resistance genes that have been cloned to date are from this gene family. Here, we describe a collection of peanut NBS–LRR resistance gene candidates (RGCs) isolated from peanut ( Arachis) species by mining Gene Bank data base. NBS–LRR sequences assembled into TIR-NBS-LRR (75.4%) and non-TIR-NBS-LRR (24.6%) subfamilies. Total of 20 distinct clades were identified and showed a high level of sequence divergence within TIR-NBS and non-TIR-NBS subfamilies. Thirty-four primer pairs were designed from these RGC sequences and used for screening different genotypes belonging to wild and cultivated peanuts. Therefore, peanut RGC identified in this study will provide useful tools for developing DNA markers and cloning the genes for resistance to different pathogens in peanut.

Peanut improvement: production of fertile hybrids and backcross progeny between Arachis hypogaea and A. kretschmeri

There are only a few reports of successful crosses between cultivated peanut (Arachis hypogaea L., section Arachis) and wild species from sections other than section Arachis. Many of the wild Arachis species harbor important traits necessary for the improvement of peanut. For example, Arachis kretschmeri Krapov., W.C. Gregory & C.E. Simpson (section Procumbentes) can grow under water-logged conditions and has been identified as one of the few wild species of Arachis with resistance to late leaf spot (LLS) and peanut rosette disease. Peanut rosette, caused by a combination of viruses, is an economically important disease only in Africa, while LLS, caused by Cercosporidium personatum, is an important fungal disease in Asia and the Americas as well as Africa. Interspecific hybrids between A. hypogaea and A. kretschmeri were produced by applying growth regulators to pollinated pistils and hybrid plants were obtained by germinating embryos in vitro. A total of seven hybrids were produced and confirmed by Simple Sequence Repeat (SSR) analysis. All hybrids were fertile, although initially slow growing. F1 hybrids were backcrossed to A. hypogaea and all plants in the F1BC1 generation were single-seeded with a prominent beak, characteristic of A. kretschmeri, but many of the F1BC2 pods were double-seeded resembling A. hypogaea. F1BC2 plants were moderately resistant to LLS. When a large number of seeds are obtained, the progeny will be screened for resistance to both LLS and rosette disease. Thus crosses with species outside the section Arachis may not only confer disease resistance but will also broaden the genetic base of cultivated peanut.

Evaluation of the U.S. peanut mini core collection using a molecular marker for resistance to Sclerotinia minor Jagger

Cultivated peanut, the second most economically important legume crop throughout the United States and the third most important oilseed in the world, is consistently threatened by various diseases and pests. Sclerotinia minor Jagger (S. minor), the causal agent of Sclerotinia blight, is a major threat to peanut production in the Southwestern U.S., Virginia, and North Carolina and can reduce yield by up to 50% in severely infested fields. Although host plant resistance would provide the most effective solution to managing Sclerotinia blight, limited sources of resistance to the disease are available for use in breeding programs. Peanut germplasm collections are available for exploration and identification of new sources of resistance, but traditionally the process is lengthy, requiring years of field testing before those potential sources can be identified. Molecular markers associated with phenotypic traits can speed up the screening of germplasm accessions, but until recently none were available for Sclerotinia blight resistance in peanut. This study objective of this study was to characterize the US peanut mini-core collection with regards to a recently discovered molecular marker associated with Sclerotinia blight resistance. Ninety-six accessions from the collection were available and genotyped using the SSR marker and 39 total accessions from spanish, valencia, runner market types were identified as new potential sources of resistance and targeted for further evaluation in field tests for Sclerotinia blight resistance.

Identification of QTLs for pod and kernel traits in cultivated peanut by bulked segregant analysis

Bulked segregant analysis was used to identify simple sequence repeat (SSR) markers associated with pod and kernel traits in cultivated peanut, to permit rapid selection of superior quality genotypes in the breeding program. SSR markers linked to pod and kernel traits were identified in two DNA pools (high and low), which were established using selected F 2:6 recombinant individuals resulting from a cultivated cross between a runner (Tamrun OL01) and a Spanish (BSS 56) peanut. To identify quantitative trait loci (QTLs) for pod and kernel-related traits, parents were screened initially with 112 SSR primer pairs. The survey revealed 8.9% polymorphism between parents. Of ten SSR primer pairs distinguishing the parents, five (PM375, PM36, PM45, pPGPseq8D9, and Ah-041) were associated with differences between bulks for seed length, pod length, number of pods per plant, 100-seed weight, maturity, or oil content. Association was confirmed by analysis of segregation among 88 F 2:6 individuals in the RIL population. Phenotypic means associated with markers for three traits differed by more than 40%, indicating the presence of QTLs with major effects for number of pods per plant, plant weight, and pod maturity. The SSR markers can be used for marker assisted selection for quality and yield improvement in peanut. To the best of our knowledge, this is the first report on the identification of SSR markers linked to pod - and kernel- related traits in cultivated peanut.

Identification of QTLs for pod and kernel traits in cultivated peanut by bulked segregant analysis

Identification of QTLs for pod and kernel traits in cultivated peanut by bulked segregant analysis

Characterization of rust, early and late leaf spot resistance in wild and cultivated peanut germplasm

Groundnut (Arachis hypogaea) has an AB genome and is one of the most important oil crops in the world. The main constraints of crop management in Brazil are fungal diseases. Several species of the genus Arachis are resistant to pests and diseases. The objective of our experiments was to identify wild species belonging to the taxonomic section Arachis with either A or B (or " non-A" ) genomes that are resistant to early leaf spot (Cercospora arachidicola), late leaf spot (Cercosporidium personatum) and rust (Puccinia arachidis). For the identification of genotypes resistant to fungal diseases, bioassays with detached leaves were done in laboratory conditions, with artificial inoculation, a controlled temperature of 25ºC and a photoperiod of 10 h light/14 h dark, for 20-42 days, depending on the fungi species. Most of the accessions of wild species were more resistant than accessions of A. hypogaea for one, two or all three fungi species studied. Arachis monticola, considered to be a possible tetraploid ancestor or a derivative of A. hypogaea, was also more susceptible to Cercosporidium personatum and Puccinia arachidis, as compared to most of the wild species. Therefore, wild germplasm accessions of both genome types are available to be used for the introgression of resistance genes against three fungal diseases of peanut.

Utility of EST-derived SSR in cultivated peanut (Arachis hypogaea L.) and Arachis wild species.

BackgroundLack of sufficient molecular markers hinders current genetic research in peanuts (Arachis hypogaea L.). It is necessary to develop more molecular markers for potential use in peanut genetic research. With the development of peanut EST projects, a vast amount of available EST sequence data has been generated. These data offered an opportunity to identify SSR in ESTs by data mining.ResultsIn this study, we investigated 24,238 ESTs for the identification and development of SSR markers. In total, 881 SSRs were identified from 780 SSR-containing unique ESTs. On an average, one SSR was found per 7.3 kb of EST sequence with tri-nucleotide motifs (63.9%) being the most abundant followed by di- (32.7%), tetra- (1.7%), hexa- (1.0%) and penta-nucleotide (0.7%) repeat types. The top six motifs included AG/TC (27.7%), AAG/TTC (17.4%), AAT/TTA (11.9%), ACC/TGG (7.72%), ACT/TGA (7.26%) and AT/TA (6.3%). Based on the 780 SSR-containing ESTs, a total of 290 primer pairs were successfully designed and used for validation of the amplification and assessment of the polymorphism among 22 genotypes of cultivated peanuts and 16 accessions of wild species. The results showed that 251 primer pairs yielded amplification products, of which 26 and 221 primer pairs exhibited polymorphism among the cultivated and wild species examined, respectively. Two to four alleles were found in cultivated peanuts, while 3–8 alleles presented in wild species. The apparent broad polymorphism was further confirmed by cloning and sequencing of amplified alleles. Sequence analysis of selected amplified alleles revealed that allelic diversity could be attributed mainly to differences in repeat type and length in the microsatellite regions. In addition, a few single base mutations were observed in the microsatellite flanking regions.ConclusionThis study gives an insight into the frequency, type and distribution of peanut EST-SSRs and demonstrates successful development of EST-SSR markers in cultivated peanut. These EST-SSR markers could enrich the current resource of molecular markers for the peanut community and would be useful for qualitative and quantitative trait mapping, marker-assisted selection, and genetic diversity studies in cultivated peanut as well as related Arachis species. All of the 251 working primer pairs with names, motifs, repeat types, primer sequences, and alleles tested in cultivated and wild species are listed in Additional File 1.

Analysis of Gene Expression Profiles in Leaf Tissues of Cultivated Peanuts and Development of EST-SSR Markers and Gene Discovery.

Peanut is vulnerable to a range of foliar diseases such as spotted wilt caused by Tomato spotted wilt virus (TSWV), early (Cercospora arachidicola) and late (Cercosporidium personatum) leaf spots, southern stem rot (Sclerotium rolfsii), and sclerotinia blight (Sclerotinia minor). In this study, we report the generation of 17,376 peanut expressed sequence tags (ESTs) from leaf tissues of a peanut cultivar (Tifrunner, resistant to TSWV and leaf spots) and a breeding line (GT-C20, susceptible to TSWV and leaf spots). After trimming vector and discarding low quality sequences, a total of 14,432 high-quality ESTs were selected for further analysis and deposition to GenBank. Sequence clustering resulted in 6,888 unique ESTs composed of 1,703 tentative consensus (TCs) sequences and 5185 singletons. A large number of ESTs (5717) representing genes of unknown functions were also identified. Among the unique sequences, there were 856 EST-SSRs identified. A total of 290 new EST-based SSR markers were developed and examined for amplification and polymorphism in cultivated peanut and wild species. Resequencing information of selected amplified alleles revealed that allelic diversity could be attributed mainly to differences in repeat type and length in the SSR regions. In addition, a few additional INDEL mutations and substitutions were observed in the regions flanking the microsatellite regions. In addition, some defense-related transcripts were also identified, such as putative oxalate oxidase (EU024476) and NBS-LRR domains. EST data in this study have provided a new source of information for gene discovery and development of SSR markers in cultivated peanut. A total of 16931 ESTs have been deposited to the NCBI GenBank database with accession numbers ES751523 to ES768453.

A linkage map for the B-genome of Arachis (Fabaceae) and its synteny to the A-genome

BackgroundArachis hypogaea (peanut) is an important crop worldwide, being mostly used for edible oil production, direct consumption and animal feed. Cultivated peanut is an allotetraploid species with two different genome components, A and B. Genetic linkage maps can greatly assist molecular breeding and genomic studies. However, the development of linkage maps for A. hypogaea is difficult because it has very low levels of polymorphism. This can be overcome by the utilization of wild species of Arachis, which present the A- and B-genomes in the diploid state, and show high levels of genetic variability.ResultsIn this work, we constructed a B-genome linkage map, which will complement the previously published map for the A-genome of Arachis, and produced an entire framework for the tetraploid genome. This map is based on an F2 population of 93 individuals obtained from the cross between the diploid A. ipaënsis (K30076) and the closely related A. magna (K30097), the former species being the most probable B genome donor to cultivated peanut. In spite of being classified as different species, the parents showed high crossability and relatively low polymorphism (22.3%), compared to other interspecific crosses. The map has 10 linkage groups, with 149 loci spanning a total map distance of 1,294 cM. The microsatellite markers utilized, developed for other Arachis species, showed high transferability (81.7%). Segregation distortion was 21.5%. This B-genome map was compared to the A-genome map using 51 common markers, revealing a high degree of synteny between both genomes.ConclusionThe development of genetic maps for Arachis diploid wild species with A- and B-genomes effectively provides a genetic map for the tetraploid cultivated peanut in two separate diploid components and is a significant advance towards the construction of a transferable reference map for Arachis. Additionally, we were able to identify affinities of some Arachis linkage groups with Medicago truncatula, which will allow the transfer of information from the nearly-complete genome sequences of this model legume to the peanut crop.

Acid Components in Arachis Species and Interspecies Hybrids with High Oleic and Low Palmitic Acids

Fatty acids of 87 wild Arachis accessions and 113 interspecific hybrid derivatives were tested to investigate their variations and breeding potential for improving fatty acid content in cultivated peanut ( A. hypogaea L.). Compared with cultivated peanut, the wild species showed similar palmitic content, lower stearic and oleic contents, and higher linoleic content. The components of fatty acids varied obviously across species or accessions in the same species. The lowest content of palmitic acid was in A. rigonii; the highest oleic acid content in A. pusilla and A. duranensis, and the highest linoleic acid content in A. batizoco. Two accessions coded as 19-6 ( A. duranensis) and 23-1 ( Arachis sp.) were identified with oleic content higher than 60%. Seven species were in high content of linoleic acid (>40%), including accession 11-4 ( A. rigonii) with 48% of linoleic acid content that was the highest record. The interspecific hybrids had transgressive segregation in most fatty acids. There were significant variations in contents of palmitic, stearic, oleic, and linoleic acids among the hybrid lines. Six hybrids were obtained with oleic acid content higher than 64% and palmitic acid content less than 8.5%, such as yz8913-8 with oleic acid content of 67.8% (30 percent points higher than its parents) and palmitic acid content of 7.6%. Analysis based on sequence-related amplified polymorphism (SRAP) showed that new bands were present in the 6 hybrids. According to pod yield per plant, hundred-pod weight, hundred-seed weight, growth duration, main stem height, and total branch number, hybrids yz8818-16, yz903-1-59, yz903-2-40, and y8913-8 were of great value in breeding programs.

Identification of candidate genome regions controlling disease resistance in Arachis

BackgroundWorldwide, diseases are important reducers of peanut (Arachis hypogaea) yield. Sources of resistance against many diseases are available in cultivated peanut genotypes, although often not in farmer preferred varieties. Wild species generally harbor greater levels of resistance and even apparent immunity, although the linkage of agronomically un-adapted wild alleles with wild disease resistance genes is inevitable. Marker-assisted selection has the potential to facilitate the combination of both cultivated and wild resistance loci with agronomically adapted alleles. However, in peanut there is an almost complete lack of knowledge of the regions of the Arachis genome that control disease resistance.ResultsIn this work we identified candidate genome regions that control disease resistance. For this we placed candidate disease resistance genes and QTLs against late leaf spot disease on the genetic map of the A-genome of Arachis, which is based on microsatellite markers and legume anchor markers. These marker types are transferable within the genus Arachis and to other legumes respectively, enabling this map to be aligned to other Arachis maps and to maps of other legume crops including those with sequenced genomes. In total, 34 sequence-confirmed candidate disease resistance genes and five QTLs were mapped.ConclusionCandidate genes and QTLs were distributed on all linkage groups except for the smallest, but the distribution was not even. Groupings of candidate genes and QTLs for late leaf spot resistance were apparent on the upper region of linkage group 4 and the lower region of linkage group 2, indicating that these regions are likely to control disease resistance.

Construction of Genetic Linkage Map Based on SSR Markers in Peanut ( Arachis hypogaea L.)

Molecular genetic maps of crop species can be used in a variety of ways in breeding and genomic research such as identification and mapping of genes and quantitative trait loci (QTLs) for morphological, physiological and economic traits of crop species. However, a comprehensive genetic linkage map for cultivated peanut has not yet been developed due to the extremely low frequency of DNA polymorphism in cultivated peanut. In this study, 142 recombinant inbred lines (RILs) derived from a cross between Yueyou 13 and Zhenzhuhei were used as mapping population in peanut ( Arachis hypogaea L.). A total 652 pairs of genomic-SSR primer and 392 pairs of EST-SSR primer were used to detect the polymorphisms between the two parents. 141 SSR primer pairs, 127 genomic-SSR and 14 EST-SSR ones, which can be used to detect polymorphisms between the two parents, were selected to analyze the RILs population. Thus, a linkage genetic map which consists of 131 SSR loci in 20 linkage groups, with a coverage of 679 cM and an average of 6.12 cM of inter-maker distance was constructed. The putative functions of 12 EST-SSR markers located on the map were analyzed. Eleven showed homology to gene sequences deposited in GenBank. This is the first report of construction of a comprehensive genetic map with SSR markers in peanut ( Arachis hypogaea L.). The map presented here will provide a genetic framework for mapping the qualitative and quantitative trait in peanut.

Peanut gene expression profiling in developing seeds at different reproduction stages during Aspergillus parasiticusinfection

BackgroundPeanut (Arachis hypogaea L.) is an important crop economically and nutritionally, and is one of the most susceptible host crops to colonization of Aspergillus parasiticus and subsequent aflatoxin contamination. Knowledge from molecular genetic studies could help to devise strategies in alleviating this problem; however, few peanut DNA sequences are available in the public database. In order to understand the molecular basis of host resistance to aflatoxin contamination, a large-scale project was conducted to generate expressed sequence tags (ESTs) from developing seeds to identify resistance-related genes involved in defense response against Aspergillus infection and subsequent aflatoxin contamination.ResultsWe constructed six different cDNA libraries derived from developing peanut seeds at three reproduction stages (R5, R6 and R7) from a resistant and a susceptible cultivated peanut genotypes, 'Tifrunner' (susceptible to Aspergillus infection with higher aflatoxin contamination and resistant to TSWV) and 'GT-C20' (resistant to Aspergillus with reduced aflatoxin contamination and susceptible to TSWV). The developing peanut seed tissues were challenged by A. parasiticus and drought stress in the field. A total of 24,192 randomly selected cDNA clones from six libraries were sequenced. After removing vector sequences and quality trimming, 21,777 high-quality EST sequences were generated. Sequence clustering and assembling resulted in 8,689 unique EST sequences with 1,741 tentative consensus EST sequences (TCs) and 6,948 singleton ESTs. Functional classification was performed according to MIPS functional catalogue criteria. The unique EST sequences were divided into twenty-two categories. A similarity search against the non-redundant protein database available from NCBI indicated that 84.78% of total ESTs showed significant similarity to known proteins, of which 165 genes had been previously reported in peanuts. There were differences in overall expression patterns in different libraries and genotypes. A number of sequences were expressed throughout all of the libraries, representing constitutive expressed sequences. In order to identify resistance-related genes with significantly differential expression, a statistical analysis to estimate the relative abundance (R) was used to compare the relative abundance of each gene transcripts in each cDNA library. Thirty six and forty seven unique EST sequences with threshold of R > 4 from libraries of 'GT-C20' and 'Tifrunner', respectively, were selected for examination of temporal gene expression patterns according to EST frequencies. Nine and eight resistance-related genes with significant up-regulation were obtained in 'GT-C20' and 'Tifrunner' libraries, respectively. Among them, three genes were common in both genotypes. Furthermore, a comparison of our EST sequences with other plant sequences in the TIGR Gene Indices libraries showed that the percentage of peanut EST matched to Arabidopsis thaliana, maize (Zea mays), Medicago truncatula, rapeseed (Brassica napus), rice (Oryza sativa), soybean (Glycine max) and wheat (Triticum aestivum) ESTs ranged from 33.84% to 79.46% with the sequence identity ≥ 80%. These results revealed that peanut ESTs are more closely related to legume species than to cereal crops, and more homologous to dicot than to monocot plant species.ConclusionThe developed ESTs can be used to discover novel sequences or genes, to identify resistance-related genes and to detect the differences among alleles or markers between these resistant and susceptible peanut genotypes. Additionally, this large collection of cultivated peanut EST sequences will make it possible to construct microarrays for gene expression studies and for further characterization of host resistance mechanisms. It will be a valuable genomic resource for the peanut community. The 21,777 ESTs have been deposited to the NCBI GenBank database with accession numbers ES702769 to ES724546.

Inheritance of a One-Seeded Pod Trait in Peanut

Normally, the cultivated peanut (Arachis hypogaea L.) has predominantly 2 seeds per pod or more. Two seeds per pod are predominantly found in A. hypogaea L. subsp. hypogaea var. hypogaea (the botanical classification of the US runner and virginia market types) and in subsp. fastigiata var. vulgaris (the US spanish market type); whereas, predominantly 3 or more seeds per pod are found in subsp. fastigiata vars. fastigiata (the US valencia market type), peruviana (not marketed in the United States), and aequatoriana (not marketed in the United States), and in subsp. hypogaea var. hirsuta (not marketed in the United States). However, recently, predominantly 1 seed per pod selections were found within a Georgia cross population. Crosses involving the 1-seeded pod selection were made to determine its inheritance. The F(1), F(2), and F(3) data indicated that any 2 of 3 duplicate recessive genes designated, osp(1), osp(2), and osp(3), control the 1-seeded pod trait in peanut.

BAC libraries construction from the ancestral diploid genomes of the allotetraploid cultivated peanut

BackgroundCultivated peanut, Arachis hypogaea is an allotetraploid of recent origin, with an AABB genome. In common with many other polyploids, it seems that a severe genetic bottle-neck was imposed at the species origin, via hybridisation of two wild species and spontaneous chromosome duplication. Therefore, the study of the genome of peanut is hampered both by the crop's low genetic diversity and its polyploidy. In contrast to cultivated peanut, most wild Arachis species are diploid with high genetic diversity. The study of diploid Arachis genomes is therefore attractive, both to simplify the construction of genetic and physical maps, and for the isolation and characterization of wild alleles. The most probable wild ancestors of cultivated peanut are A. duranensis and A. ipaënsis with genome types AA and BB respectively.ResultsWe constructed and characterized two large-insert libraries in Bacterial Artificial Chromosome (BAC) vector, one for each of the diploid ancestral species. The libraries (AA and BB) are respectively c. 7.4 and c. 5.3 genome equivalents with low organelle contamination and average insert sizes of 110 and 100 kb. Both libraries were used for the isolation of clones containing genetically mapped legume anchor markers (single copy genes), and resistance gene analogues.ConclusionThese diploid BAC libraries are important tools for the isolation of wild alleles conferring resistances to biotic stresses, comparisons of orthologous regions of the AA and BB genomes with each other and with other legume species, and will facilitate the construction of a physical map.

Genetic relationships among Arachis hypogaea L. (AABB) and diploid Arachis species with AA and BB genomes

The cultivated peanut (Arachis hypogaea L.) is an allotetraploid, with two types of genomes, classified as AA and BB, according to cytogenetic characters. Similar genomes to those of A. hypogaea are found in the wild diploid species of section Arachis, which is one of the nine Arachis sections. The wild species have resistances to pests and diseases that affect the cultivated peanut and are a potential source of genes to increase the resistance levels in peanut. The aim of this study was to analyze the genetic variability within AA and BB genome species and to evaluate how they are related to each other and to A. hypogaea, using RAPD markers. Eighty-seven polymorphic bands amplified by ten 10-mer primers were analyzed. The species were divided into two major groups, and the AA and the BB genome species were, in general, separated from each other. The results showed that high variation is available within species that have genomes similar to the AA and the BB genomes of A. hypogaea.

Genetic Diversity in Cultivated Groundnut Based on SSR Markers

Peanut ( Arachis hypogaea L.) is an important source crop for edible oil and protein. It is important to identify the genetic diversity of peanut genetic resources for cultivar development and evaluation of peanut accessions. Thirty-four SSR markers were used to assess the genetic variation of four sets of twenty-four accessions each from the four botanical varieties of the cultivated peanut. Among the tested accessions, ten to sixteen pairs of SSR primers showed polymorphisms. The maximum differentiation index, which was defined as the degree of genetic differentiation, was as high as 0.992 in the tested accessions. Each accession could be discriminated by a specific set of polymorphic SSR primers, and the intra-variety genetic distance was determined among accessions, with an average of 0.59 in var. fastigiata 0.46 in var. hypogaea 0.38 in var. vulgaris and 0.17 in var. hirsuta. Dendrogrames based on genetic distances were constructed for the four botanical varieties, which revealed the existence of different clusters. It was concluded that there was abundant intra-variety SSR polymorphism, and with more and more SSR markers being developed, the intrinsic genetic diversity would be detected and the development of genetic map and marker-assisted selection for cultivated peanut would be feasible.

ESTs from a wild Arachis species for gene discovery and marker development.

BackgroundDue to its origin, peanut has a very narrow genetic background. Wild relatives can be a source of genetic variability for cultivated peanut. In this study, the transcriptome of the wild species Arachis stenosperma accession V10309 was analyzed.ResultsESTs were produced from four cDNA libraries of RNAs extracted from leaves and roots of A. stenosperma. Randomly selected cDNA clones were sequenced to generate 8,785 ESTs, of which 6,264 (71.3%) had high quality, with 3,500 clusters: 963 contigs and 2537 singlets. Only 55.9% matched homologous sequences of known genes. ESTs were classified into 23 different categories according to putative protein functions. Numerous sequences related to disease resistance, drought tolerance and human health were identified. Two hundred and six microsatellites were found and markers have been developed for 188 of these. The microsatellite profile was analyzed and compared to other transcribed and genomic sequence data.ConclusionThis is, to date, the first report on the analysis of transcriptome of a wild relative of peanut. The ESTs produced in this study are a valuable resource for gene discovery, the characterization of new wild alleles, and for marker development. The ESTs were released in the [GenBank:EH041934 to EH048197].

Molecular analysis of Arachis interspecific hybrids

Incorporation of genetic resistance against several biotic stresses that plague cultivated peanut, Arachis hypogaea (2n = 4x = 40), is an ideal option to develop disease resistant and ecologically safe peanut varieties. The primary gene pool of peanut contains many diploid wild species (2n = 2x = 20) of Arachis, which have high levels of disease and insect resistances. However, transfer of resistant genes from these species into A. hypogaea is difficult due to ploidy level differences and genomic incompatibilities. This study was conducted to monitor alien germplasm transmission, using Random Amplified Polymorphic DNA (RAPD) markers, from two diploid wild species, A. cardenasii and A. batizocoi, into A. hypogaea. Triploid interspecific hybrids were produced by crossing two A. hypogaea cultivars (NC 6 and Argentine) with the two species and by colchicine-treating vegetative meristems, fertility was restored at the hexaploid (C(o)) level in the four hybrids. Hexaploids were allowed to self-pollinate for four generations, each referred to as a cycle (C1, C2, C3, and C4). At each cycle, a backcross was made with the respective A. hypogaea cultivar as the maternal parent and only lineages tracing back to a single hexaploid hybrid were used for RAPD analysis. Analysis of mapped, species-specific RAPD markers in BC1F1 to BC1F3 hybrids indicated that alien germplasm retention decreased every generation of inbreeding, especially in Argentine and in A. batizocoi crosses. A similar trend was also observed for every cycle in BC1F2 and BC1F3 families, possibly, due to the loss of alien chromosomes following selfing of hexaploids. RAPD marker analysis of 40-chromosome interspecific hybrid derivatives from the four crosses supported previous reports that reciprocal recombination and/or translocations are the predominant mechanisms for exchange of chromosomal segments. No evidence was found for preferential transfer of alien chromosomal regions to specific linkage groups. The implications for developing disease resistant peanut breeding lines are discussed in light of these findings.