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To test the hypothesis that the cultivated peanut species possesses almost no molecular variability, we sequenced a diverse panel of 22 Arachis accessions representing Arachis hypogaea botanical classes, A-, B-, and K- genome diploids, a synthetic amphidiploid, and a tetraploid wild species. RNASeq was performed on pools of three tissues, and de novo assembly was performed. Realignment of individual accession reads to transcripts of the cultivar OLin identified 306,820 biallelic SNPs. Among 10 naturally occurring tetraploid accessions, 40,382 unique homozygous SNPs were identified in 14,719 contigs. In eight diploid accessions, 291,115 unique SNPs were identified in 26,320 contigs. The average SNP rate among the 10 cultivated tetraploids was 0.5, and among eight diploids was 9.2 per 1000 bp. Diversity analysis indicated grouping of diploids according to genome classification, and cultivated tetraploids by subspecies. Cluster analysis of variants indicated that sequences of B genome species were the most similar to the tetraploids, and the next closest diploid accession belonged to the A genome species. A subset of 66 SNPs selected from the dataset was validated; of 782 SNP calls, 636 (81.32%) were confirmed using an allele-specific discrimination assay. We conclude that substantial genetic variability exists among wild species. Additionally, significant but lesser variability at the molecular level occurs among accessions of the cultivated species. This survey is the first to report significant SNP level diversity among transcripts, and may explain some of the phenotypic differences observed in germplasm surveys. Understanding SNP variants in the Arachis accessions will benefit in developing markers for selection.

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Peanut is the second-most important legum e grown worldwide. Cultivated peanut is a disomic tetraploid, 2n-4 x -40, with limited genetic diversity due to a genetic bottleneck in formation o f the polyploid from ancestors A. duranensis and A. ipaensis. Consequently, resistance_to biotic stresses is limited in the cultigen; however, w ild species possess strong resistances. Transfer o f these re sistances is hindered by differences o f ploidy, but production o f synthetic amphidiploids, coupled with use o f m olecular markers, enables efficient gene transfer. Marker maps have been made from interspecific crosses, and SSR-based maps from cultivated parents have been developed recently. At least 410 resistance gene analogues have been identified. The first markers for biotic stress tolerance were for root-knot nematode resistance and introgressed from one A. cardenasii chromosome. These and improved markers have been used for marker-assisted backcrossing, contributing to release o f three cultivars. Additional QTLs have been identified since.Early and late leafspots cause significant yield losses worldwide, and resistance depends on multiple genes. U sing interspecific populations, five resistance QTLs for early leafspot were identified using greenhouse inoculations, and five QTLs for late leafspot were identified using detached leaf assays. U sing cultivated species populations, 28 QTLs were identified for LLS resistance; all but one w ere minor QTLs; the major QTL was donated by an interspecific introgression line parent. Rust often occurs alongside leafspots, and rust resistance was characterized as one major QTL, plus several smaller QTLs. Marker-assisted backcrossing o f this major QTL has been performed into different populations. QTLs for resistance to other biotic stresses have been identified, namely to groundnut rosette virus, Sclerotinia blight, afiatoxin contamination, aphids, and tomato spotted w ilt virus. Marker-assisted breeding is still in early stages, and develop ment o f more rapid and inexpensive markers from transcriptome and genom e sequencing is expected to accelerate progress.

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