Abundant Microsatellite Diversity and Oil Content in Wild Arachis Species

The peanut (Arachis hypogaea) is an important oil crop. Breeding for high oil content is becoming increasingly important. Wild Arachis species have been reported to harbor genes for many valuable traits that may enable the improvement of cultivated Arachis hypogaea, such as resistance to pests and disease. However, only limited information is available on variation in oil content. In the present study, a collection of 72 wild Arachis accessions representing 19 species and 3 cultivated peanut accessions were genotyped using 136 genome-wide SSR markers and phenotyped for oil content over three growing seasons. The wild Arachis accessions showed abundant diversity across the 19 species. A. duranensis exhibited the highest diversity, with a Shannon-Weaver diversity index of 0.35. A total of 129 unique alleles were detected in the species studied. A. rigonii exhibited the largest number of unique alleles (75), indicating that this species is highly differentiated. AMOVA and genetic distance analyses confirmed the genetic differentiation between the wild Arachis species. The majority of SSR alleles were detected exclusively in the wild species and not in A. hypogaea, indicating that directional selection or the hitchhiking effect has played an important role in the domestication of the cultivated peanut. The 75 accessions were grouped into three clusters based on population structure and phylogenic analysis, consistent with their taxonomic sections, species and genome types. A. villosa and A. batizocoi were grouped with A. hypogaea, suggesting the close relationship between these two diploid wild species and the cultivated peanut. Considerable phenotypic variation in oil content was observed among different sections and species. Nine alleles were identified as associated with oil content based on association analysis, of these, three alleles were associated with higher oil content but were absent in the cultivated peanut. The results demonstrated that there is great potential to increase the oil content in A. hypogaea by using the wild Arachis germplasm.     

Production of peanut hybrid seeds in an intersectional cross through post-pollination treatment of flower bases with plant growth regulators

To broaden the narrow gene base of the cultivated peanut and utilize sources of resistance to abiotic/biotic stresses, quality-related traits and high yielding factors, much attention has been paid to wild relatives of the oilseed crop since 1980s. Species outside section Arachis are cross-incompatible with A. hypogaea L.; even some of the species from section Arachis, which are supposed to be compatible, may also encounter obstacles when crossed with the peanut cultigen. The objective of the present communication is to study the effects of simple hormone treatment on production of true incompatible hybrids in peanut to replace the tedious and lengthy in vitro embryo rescue procedures currently in use. In the incompatible cross, A. hypogaea cv Qunyu 101?×?A. paraguariensis, post-pollination application of a hormone aqueous solution (IAA 4?mg/L?+?GA 2?mg/L) at flower bases resulted in 42 seeds, of which 26 were identified as true hybrids by allele-specific PCR for FAD2A genotyping. As a desirable alternative to in vitro embryo/ovules/peg culture, the present method may facilitate the utilization of wild species in peanut breeding. The method described here for peanut may be of reference to other crop plants where embryo abortion is also a problem.     

Identification of Fungus Resistant Wild Accessions and Interspecific Hybrids of the Genus Arachis

Peanut, Arachis hypogaea L., is a protein-rich species consumed worldwide. A key improvement to peanut culture involves the development of cultivars that resist fungal diseases such as rust, leaf spot and scab. Over three years, we evaluated fungal resistance under field conditions of 43 wild accessions and three interspecific hybrids of the genus Arachis, as well as six A. hypogaea genotypes. In the first year, we evaluated resistance to early and late leaf spot, rust and scab. In the second and third years, we evaluated the 18 wild species with the best resistance scores and control cultivar IAC Caiapó for resistance to leaf spot and rust. All wild accessions displayed greater resistance than A. hypogaea but differed in their degree of resistance, even within the same species. We found accessions with as good as or better resistance than A. cardenasii, including: A. stenosperma (V15076 and Sv 3712), A. kuhlmannii (V 6413), A. kempff-mercadoi (V 13250), A. hoehnei (KG 30006), and A. helodes (V 6325). Amphidiploids and hybrids of A. hypogaea behaved similarly to wild species. An additional four accessions deserve further evaluation: A. magna (V 13751 and KG 30097) and A. gregoryi (V 14767 and V 14957). Although they did not display as strong resistance as the accessions cited above, they belong to the B genome type that is crucial to resistance gene introgression and pyramidization in A. hypogaea.     

Profiling microRNA expression in Arabidopsis pollen using microRNA array and real-time PCR

Background

Arachis 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.

Results

In 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.

Conclusion

The 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.     

Utilization of wild relatives in the genetic improvement of Arachis hypogaea L.

Summary Synthetic amphidiploids were established in 32 combinations involving 8 diploid wild species representing both A and B genomes of section Arachis. Bivalent and multivalent associations in the amphidiploids of 7 A genome species confirm that these species have identical genomes. Contrastingly, high bivalent frequencies in amphidiploids involving the A and B genome species suggest that A. batizocoi has a distinct B genome that is partially homologous to the other genome A represented in the rest of the species. Crossability, chromosome pairing and pollen and pod fertility in hybrids between A. hypogaea and amphidiploids have revealed that these amphidiploids can be used as a genetic bridge for the transfer of genes from the wild species into the cultivated groundnut.Submitted as Journal Article No. 530 by International Crops Research Institute for the Semi-Arid Tropics (ICRISAT)     

Early Reproductive Ontogeny in Interspecific Crosses of Arachis hypogaea and Section Arachis Species

Wild Arachis species have been recognized as sources of resistanceto pests and pathogens that infect A. hypogaea L. and causesubstantial yield losses. However, utilization of these geneticresources for crop improvement has been difficult. This studywas conducted to (a) understand the processes of early embryogrowth and development in four Arachis species, two A. hypogaeacultivars and their hybrids and (b) identify parental compatibilitiesin reciprocal crosses of A. hypogaea. The results indicatedthat delayed fertilization beyond 24 h, coupled with slow proembryogrowth, leads to embryo abortion in many interspecific crosses.For example, in female A. cardenasii crosses, lack of or delayedfertilization leads to failure to obtain hybrids. When A. batizocoiwas used as a female parent, delayed fertilization and the inabilityof quiescent proembryos to resume growth after soil penetrationcaused abortion. Embryos of A. hypogaea x A. glandulifera crossesdeveloped normally during the first 21 d after fertilization,but then aborted at a later time. In this study, A. hypogaeawas always a better female parent than the wild Arachis species.Increasing the number of pollinations per cross, using the cultivatedspecies as the female parent, utilizing different A. hypogaeavarieties, and embryo rescue techniques are suggested to improvethe probability of obtaining interspecific hybrids in Arachis.Copyright1995, 1999 Academic Press Peanut, interspecific hybrids, Arachis, wild species, incompatibilities     

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, C_3, 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.     

Introgression of homeologous quantitative trait loci (QTLs) for resistance to the root-knot nematode [Meloidogyne arenaria (Neal) Chitwood] in an advanced backcross-QTL population of peanut (Arachis hypogaea L.)

Resistance to root-knot nematodes [Meloidogyne arenaria (Neal) Chitwood] is needed for cultivation of peanut in major peanut-growing areas, but significant resistance is lacking in the cultivated species (Arachis hypogaea L.). Markers to two closely-linked genes introgressed from wild relatives of peanut have been identified previously, but phenotypic evidence for the presence of additional genes in wild species and introgression lines has eluded quantitative trait locus (QTL) identification. Here, to improve sensitivity to small-effect QTLs, an advanced backcross population from a cross between a Florunner component line and the synthetic amphidiploid TxAG-6 [Arachis batizocoi × (A. cardenasii × A. diogoi)]^4× was screened for response to root-knot nematode infection. Composite interval mapping results suggested a total of seven QTLs plus three putative QTLs. These included the known major resistance gene plus a second QTL on LG1, and a potentially homeologous B-genome QTL on LG11. Additional potential homeologs were identified on linkage group (LG) 8 and LG18, plus a QTL on LG9.2 and putative QTLs on LG9.1 and 19. A QTL on LG15 had no inferred resistance-associated homeolog. Contrary to expectation, two introgressed QTLs were associated with susceptibility, and QTLs at some homeologous loci were found to confer opposite phenotypic responses. Long-term functional conservation accompanied by rapid generation of functionally divergent alleles may be a singular feature of NBS-LRR resistance gene clusters, contributing to the richness of resistance alleles available in wild relatives of crops. The significance for peanut evolution and breeding is discussed.     

Origins of resistances to rust and late leaf spot in peanut (Arachis hypogaea,Fabaceae)

The cultivated peanut (Arachis hypogaea, Fabaceae) is believed to have originated along the eastern slopes of the Andes in Bolivia and northern Argentina. The crop is now grown throughout tropical and warm temperate regions. Among diseases attacking peanuts, rust caused byPuccinia arachidis and late leaf spot caused byPhaeoisariopsis personata are the most important and destructive on a worldwide scale. Both pathogens, restricted in host range to Arachis, probably originated and coevolved in South America along with their hosts. In recent years there has been much emphasis on screening of peanut germplasm for resistance to these diseases. At the International Crops Research Institute for the Semi-Arid Tropics (ICRISA T), India, some 10,000 peanut germplasm accessions were screened for resistance to rust and late leaf spot during 1977–1985 and sources of resistance indentified for either or both pathogens. Of the resistant genotypes, about 87% belonged to A. hypogaea var.fastigiata and 13% to var.hypogaea; 84% originated in South America or had South American connections. A high percentage (75%) had their origin in Peru (believed to be a secondary gene center for var.hirsuta and var.fastigiata,), suggesting that resistance to rust and late leaf spot diseases might have evolved in that country.     

Jasmonate-dependent plant defense restricts thrips performance and preference

Background

Worldwide, 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.

Results

In 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.

Conclusion

Candidate 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.     

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.     

Use of single-primer DNA amplifications in genetic studies of peanut (Arachis hypogaea L.)

A recent approach to detecting genetic polymorphism involves the amplification of genomic DNA using single primers of arbitrary sequence. When separated electrophoretically in agarose gels, the amplification products give banding patterns that can be scored for genetic variation. The objective of this research was to apply these techniques to cultivated peanut (Arachis hypogaea L.) and related wild species to determine whether such an approach would be feasible for the construction of a genetic linkage map in peanut or for systematic studies of the genus. Two peanut cultivars, 25 unadapted germplasm lines of A. hypogaea, the wild allotetraploid progenitor of cultivated peanut (A. monticola), A. glabrata (a tetraploid species from section Rhizomatosae), and 29 diploid wild species of Arachis were evaluated for variability using primers of arbitrary sequence to amplify segments of genomic DNA. No variation in banding pattern was observed among the cultivars and germplasm lines of A. hypogaea, whereas the wild Arachis species were uniquely identified with most primers tested. Bands were scored (+/–) in the wild species and the PAUP computer program for phylogenetic analysis and the HyperRFLP program for genetic distance analysis were used to generate dendrograms showing genetic relationships among the diploid Arachis species evaluated. The two analyses produced nearly identical dendrograms of species relationships. In addition, approximately 100 F₂ progeny from each of two interspecific crosses were evaluated for segregation of banding patterns. Although normal segregation was observed among the F₂ progeny from both crosses, banding patterns were quite complex and undesirable for use in genetic mapping. The dominant behavior of the markers prevented the differentiation of heterozygotes from homozygotes with certainty, limiting the usefulness of arbitrary primer amplification products as markers in the construction of a genetic linkage map in peanut.     

A microsatellite-based, gene-rich linkage map for the AA genome of Arachis (Fabaceae)

Cultivated peanut (Arachis hypogaea) is an important crop, widely grown in tropical and subtropical regions of the world. It is highly susceptible to several biotic and abiotic stresses to which wild species are resistant. As a first step towards the introgression of these resistance genes into cultivated peanut, a linkage map based on microsatellite markers was constructed, using an F₂ population obtained from a cross between two diploid wild species with AA genome (A. duranensis and A. stenosperma). A total of 271 new microsatellite markers were developed in the present study from SSR-enriched genomic libraries, expressed sequence tags (ESTs), and by “data-mining” sequences available in GenBank. Of these, 66 were polymorphic for cultivated peanut. The 271 new markers plus another 162 published for peanut were screened against both progenitors and 204 of these (47.1%) were polymorphic, with 170 codominant and 34 dominant markers. The 80 codominant markers segregating 1:2:1 (P<0.05) were initially used to establish the linkage groups. Distorted and dominant markers were subsequently included in the map. The resulting linkage map consists of 11 linkage groups covering 1,230.89 cM of total map distance, with an average distance of 7.24 cM between markers. This is the first microsatellite-based map published for Arachis, and the first map based on sequences that are all currently publicly available. Because most markers used were derived from ESTs and genomic libraries made using methylation-sensitive restriction enzymes, about one-third of the mapped markers are genic. Linkage group ordering is being validated in other mapping populations, with the aim of constructing a transferable reference map for Arachis.Electronic supplementary material is available for this at     

Comparison of Embryo Development in Wild and Cultivated Arachis Species

Embryo development following selfing was investigated in twowild diploid peanut species, Arachis batizocoi Krap. et Greg.(coll. K 9484) (2n = 20) and A. duranensis Krap. et Greg. nom.nud. (coll. K 7988) (2n = 20), and one cultivated tetraploidspecies, A. hypogaea L. NC-Ac 18000 (2n = 40). Rates of pegelongation and sequences of embryo development for each specieswere compared. Peg elongation rates were similar for the twowild species, but for A. hypogaea it was only one-third to one-halfthat of the diploid species. Embryos in A. hypogaea showed slightlymore rapid cell division than in the wild species. The observedvariation in reproductive development between the wild and cultivatedspecies indicate that different control mechanisms may governdevelopment in the different species and may be at least partiallyresponsible for failure to produce viable interspecific hybridsat various ploidy levels. The observations are also importantfor determining the time at which embryos of different speciesof Arachis will reach the appropriate stage of development forsuccessful culture on an artificial medium during embryo rescueprocedures. Peanut, Arachis hypogaea, wild species, embryo, peg     

A Study of Gene Expression in the Nematode Resistant Wild Peanut Relative, Arachis stenosperma, in Response to Challenge with Meloidogyne arenaria

Peanut (Arachis hypogaea) is amongst the most important legume crops in the world. One of its main yield constraints is the root-knot nematode Meloidogyne arenaria. A number of wild Arachis species, including A. stenosperma, are resistant to nematodes, and are a potential source of new resistance alleles for cultivated peanut. Using in silico subtraction of ESTs and macroarray analysis, we identified genes differentially expressed in A. stenosperma roots during its resistance response to M. arenaria. The three most differentially expressed genes [Auxin Repressed Protein (AsARP), Cytokinin Oxidase (AsCKX) and Metallothionein Type 2 (AsMET2)] were further analyzed using northern-blot and showed distinct expression profiles in the resistant A. stenosperma and susceptible A. hypogaea, both after, and sometimes even before, challenge with nematodes. Of the three most differentially expressed genes, AsARP and AsCKX are potentially involved in plant hormonal balance, and AsMET2 may be related to the reactive oxygen reaction triggered by the hypersensitive response (HR).     

Tetrasomic Recombination Is Surprisingly Frequent in Allotetraploid Arachis

Arachis hypogaea L. (cultivated peanut) is an allotetraploid (2n = 4x = 40) with an AABB genome type. Based on cytogenetic studies it has been assumed that peanut and wild-derived induced AABB allotetraploids have classic allotetraploid genetic behavior with diploid-like disomic recombination only between homologous chromosomes, at the exclusion of recombination between homeologous chromosomes. Using this assumption, numerous linkage map and quantitative trait loci studies have been carried out. Here, with a systematic analysis of genotyping and gene expression data, we show that this assumption is not entirely valid. In fact, autotetraploid-like tetrasomic recombination is surprisingly frequent in recombinant inbred lines generated from a cross of cultivated peanut and an induced allotetraploid derived from peanut’s most probable ancestral species. We suggest that a better, more predictive genetic model for peanut is that of a “segmental allotetraploid” with partly disomic, partly tetrasomic genetic behavior. This intermediate genetic behavior has probably had a previously overseen, but significant, impact on the genome and genetics of cultivated peanut.