Abstract
BackgroundUntil recently, only a small number of low- and mid-throughput methods have been used for single nucleotide polymorphism (SNP) discovery and genotyping in grapevine (Vitis vinifera L.). However, following completion of the sequence of the highly heterozygous genome of Pinot Noir, it has been possible to identify millions of electronic SNPs (eSNPs) thus providing a valuable source for high-throughput genotyping methods.ResultsHerein we report the first application of the SNPlex™ genotyping system in grapevine aiming at the anchoring of an eukaryotic genome. This approach combines robust SNP detection with automated assay readout and data analysis. 813 candidate eSNPs were developed from non-repetitive contigs of the assembled genome of Pinot Noir and tested in 90 progeny of Syrah × Pinot Noir cross. 563 new SNP-based markers were obtained and mapped. The efficiency rate of 69% was enhanced to 80% when multiple displacement amplification (MDA) methods were used for preparation of genomic DNA for the SNPlex assay.ConclusionUnlike other SNP genotyping methods used to investigate thousands of SNPs in a few genotypes, or a few SNPs in around a thousand genotypes, the SNPlex genotyping system represents a good compromise to investigate several hundred SNPs in a hundred or more samples simultaneously. Therefore, the use of the SNPlex assay, coupled with whole genome amplification (WGA), is a good solution for future applications in well-equipped laboratories.
Highlights
Until recently, only a small number of low- and mid-throughput methods have been used for single nucleotide polymorphism (SNP) discovery and genotyping in grapevine (Vitis viniferaL.)
RAPDs (Random Amplified Polymorphic DNAs), SNPs can be detected through non-gel-based high-throughput assays, saving both time and money [3]
Several SNP assay technologies have been developed based on various methods of allelic discrimination and detection platforms
Summary
Only a small number of low- and mid-throughput methods have been used for single nucleotide polymorphism (SNP) discovery and genotyping in grapevine (Vitis viniferaL.). In the last few years, single nucleotide polymorphisms (SNPs) have become the most popular genetic marker system in both animals and plants Their extraordinary abundance discovered in several genome sequencing projects [1], combined with recent technological improvements, makes SNP markers attractive for high-throughput use in marker-assisted breeding, EST mapping and the integration of genetic and physical maps. At present several SNP identification methods are available such as resequencing of PCR amplicons with or without pre-screening, electronic SNP (eSNP) discovery in expressed sequence tag (EST) and shotgun genomic libraries. In these latter cases, sequences may be computationally screened for polymorphisms to distinguish true polymorphisms from sequencing errors if sufficient redundancy is present [2]. We report a successful application of the SNPlex genotyping system, which provided 563 new SNP-based markers anchoring the grapevine genome for future applied research programs
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