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G4008.06: Single nucleotide polymorphism discovery, validation, and mapping in groundnut

Knapp, S.J., Ozias-Akins, P., Hoisington, D., Aruna, R. and Varshney, R.ORCID: 0000-0002-4562-9131 (2011) G4008.06: Single nucleotide polymorphism discovery, validation, and mapping in groundnut. Project Updates. CGIAR Generation Challenge Program .


1. Research activities and progress at UGA DNA marker resources have been expanded for molecular breeding applications in groundnut (Arachis hypogaea L.). The objective to enhance the infrastructure for translational genomics and molecular breeding research in groundnut has been achieved by massively parallel sequencing of 17 tetraploid genotypes which, by comparison with the reference transcriptome of ‘Tifrunner’, has provided a database for SNP discovery. Over 350 Mb of sequence from root, leaf, and pod tissues of 17 genotypes was assembled along with Sanger and Roche 454 sequences from the reference ‘Tifrunner’ transcriptome. 8486 single nucleotide polymorphisms (SNPs) were identified when the data were subjected to moderately stringent filtering to account for a SNP in at least two sequences from a genotype, allele frequency among genotypes, sequence errors at ends of reads, and proximity to neighboring SNPs or indels. An Illumina Golden Gate 1536-SNP array was designed from these 8486 candidate SNPs by prioritizing based on Illumina design score and distance from predicted intron-exon boundaries. The Golden Gate assay was used for genotyping of 80 tetraploid inbred lines, 3 amphidiploids, and several diploid accessions of Arachis. Loci could be detected for >95% of the SNP assays indicating successful design using this platform. However, SNPs between tetraploid genotypes were rare unless the tetraploid was synthetic. Nevertheless, the validated SNPs can be used to construct a smaller chip or transferred to an alternate platform for lower throughput assays. Sequence data from non-coding regions of the groundnut genome will be needed to increase the probability of finding nucleotide differences between cultivated genotypes in order to enhance the density of markers that can be used for breeding.

The SNPs identified and validated in this project can be used for genetic mapping of groundnut populations using several SNP assay platforms. The Illumina GoldenGate platform is most suitable for advanced recombinant inbred lines since the primers almost always amplified homeologous regions of the genome and intergenomic “SNPs” were detected as heterozygotes which would interfere with detection of a true heterozygote within a sub-genome. Groundnut breeding programs with the capability for molecular breeding can use the discovered markers initially in QTL linkage analyses and subsequently for marker-assisted selection.

2. Research activities at ICRISAT The SNP array was used to analyze genotypes of interest to the ICRISAT breeding program where many of the parental lines have been crossed in various combinations. These populations are at various stages of development although some are at the recombinant inbred line stage appropriate for Golden Gate analysis.

Tangible outputs delivered
• 350 Mb of 454-generated reduced representation sequence from 17 genotypes and an assembly of ~211,000 contigs from 21 genotypes, providing a reference transcriptome for tetraploid groundnut.

•Database of over 8000 SNPs identified from these genotypes.

•SNP genotypes (1536 loci) for 80 elite and exotic tetraploid groundnut lines have been generated and can guide mapping population development and analysis.

References Manish K. Pandey, Emmanuel Monyo, Peggy Ozias-Akins, Xuanquiang Liang, Patricia Guimarães, Shyam N. Nigam, Hari D. Upadhyaya, P. Janila, Baozhu Guo, Douglas R. Cook, David J. Bertioli, Richard Michelmore, Rajeev K. Varshney. Advances in Arachis genomics for peanut improvement. In preparation for Molec. Breed.

Item Type: Others
Publisher: CIMMYT
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