Transcript-specific, single-nucleotide polymorphism discovery and linkage analysis in hexaploid bread wheat (Triticum aestivum L.).

Food security is a global concern and substantial yield increases in cereal crops are required to feed the growing world population. Wheat is one of the three most important crops for human and livestock feed. However, the complexity of the genome coupled with a decline in genetic diversity within modern elite cultivars has hindered the application of marker-assisted selection (MAS) in breeding programmes. A crucial step in the successful application of MAS in breeding programmes is the development of cheap and easy to use molecular markers, such as single-nucleotide polymorphisms. To mine selected elite wheat germplasm for intervarietal single-nucleotide polymorphisms, we have used expressed sequence tags derived from public sequencing programmes and next-generation sequencing of normalized wheat complementary DNA libraries, in combination with a novel sequence alignment and assembly approach. Here, we describe the development and validation of a panel of 1114 single-nucleotide polymorphisms in hexaploid bread wheat using competitive allele-specific polymerase chain reaction genotyping technology. We report the genotyping results of these markers on 23 wheat varieties, selected to represent a broad cross-section of wheat germplasm including a number of elite UK varieties. Finally, we show that, using relatively simple technology, it is possible to rapidly generate a linkage map containing several hundred single-nucleotide polymorphism markers in the doubled haploid mapping population of Avalon × Cadenza.

Multiplex single nucleotide polymorphism (SNP)-based genotyping in allohexaploid wheat using padlock probes.

Single nucleotide polymorphisms are the most common polymorphism in plant and animal genomes and, as such, are the logical choice for marker-assisted selection. However, many plants are also polyploid, and marker-assisted selection can be complicated by the presence of highly similar, but non-allelic, homoeologous sequences. Despite this, there is practical and academic demand for high-throughput genotyping in several polyploid crop species, such as allohexaploid wheat. In this paper, we present such a system, which utilizes public single nucleotide polymorphisms previously identified in both agronomically important genes and in randomly selected, mapped, expressed sequence tags developed by the wheat community. To achieve relatively high levels of multiplexing, we used non-amplified genomic DNA and padlock probe pairs, together with high annealing temperatures, to differentiate between similar sequences in the wheat genome. Our results suggest that padlock probes are capable of discriminating between homoeologous sequences and hence can be used to efficiently genotype wheat varieties.

Single nucleotide polymorphisms and linkage disequilibrium in sunflower.

Genetic diversity in modern sunflower (Helianthus annuus L.) cultivars (elite oilseed inbred lines) has been shaped by domestication and breeding bottlenecks and wild and exotic allele introgression(-)the former narrowing and the latter broadening genetic diversity. To assess single nucleotide polymorphism (SNP) frequencies, nucleotide diversity, and linkage disequilibrium (LD) in modern cultivars, alleles were resequenced from 81 genic loci distributed throughout the sunflower genome. DNA polymorphisms were abundant; 1078 SNPs (1/45.7 bp) and 178 insertions-deletions (INDELs) (1/277.0 bp) were identified in 49.4 kbp of DNA/genotype. SNPs were twofold more frequent in noncoding (1/32.1 bp) than coding (1/62.8 bp) sequences. Nucleotide diversity was only slightly lower in inbred lines ( = 0.0094) than wild populations ( = 0.0128). Mean haplotype diversity was 0.74. When extraploted across the genome ( approximately 3500 Mbp), sunflower was predicted to harbor at least 76.4 million common SNPs among modern cultivar alleles. LD decayed more slowly in inbred lines than wild populations (mean LD declined to 0.32 by 5.5 kbp in the former, the maximum physical distance surveyed), a difference attributed to domestication and breeding bottlenecks. SNP frequencies and LD decay are sufficient in modern sunflower cultivars for very high-density genetic mapping and high-resolution association mapping.