High-density single nucleotide polymorphism (SNP) genotyping arrays are a powerful tool for studying genomic patterns of diversity, inferring ancestral relationships between individuals in populations and studying marker-trait associations in mapping experiments. We developed a genotyping array including about 90,000 gene-associated SNPs and used it to characterize genetic variation in allohexaploid and allotetraploid wheat populations. The array includes a significant fraction of common genome-wide distributed SNPs that are represented in populations of diverse geographical origin. We used density-based spatial clustering algorithms to enable high-throughput genotype calling in complex data sets obtained for polyploid wheat. We show that these model-free clustering algorithms provide accurate genotype calling in the presence of multiple clusters including clusters with low signal intensity resulting from significant sequence divergence at the target SNP site or gene deletions. Assays that detect low-intensity clusters can provide insight into the distribution of presence-absence variation (PAV) in wheat populations. A total of 46 977 SNPs from the wheat 90K array were genetically mapped using a combination of eight mapping populations. The developed array and cluster identification algorithms provide an opportunity to infer detailed haplotype structure in polyploid wheat and will serve as an invaluable resource for diversity studies and investigating the genetic basis of trait variation in wheat.

/pdf/characterization-of-polyploid-wheat-genomic-diversity-using-2k5bjm1mv0.pdf

Characterization of polyploid wheat genomic diversity using a high-density 90 000 single nucleotide polymorphism array

An ordered draft sequence of the 17-gigabase hexaploid bread wheat (Triticum aestivum) genome has been produced by sequencing isolated chromosome arms. We have annotated 124,201 gene loci distributed nearly evenly across the homeologous chromosomes and subgenomes. Comparative gene analysis of wheat subgenomes and extant diploid and tetraploid wheat relatives showed that high sequence similarity and structural conservation are retained, with limited gene loss, after polyploidization. However, across the genomes there was evidence of dynamic gene gain, loss, and duplication since the divergence of the wheat lineages. A high degree of transcriptional autonomy and no global dominance was found for the subgenomes. These insights into the genome biology of a polyploid crop provide a springboard for faster gene isolation, rapid genetic marker development, and precise breeding to meet the needs of increasing food demand worldwide.

A chromosome-based draft sequence of the hexaploid bread wheat (Triticum aestivum) genome

Complete and accurate reference genomes and annotations provide fundamental tools for characterization of genetic and functional variation. These resources facilitate the determination of biological processes and support translation of research findings into improved and sustainable agricultural technologies. Many reference genomes for crop plants have been generated over the past decade, but these genomes are often fragmented and missing complex repeat regions. Here we report the assembly and annotation of a reference genome of maize, a genetic and agricultural model species, using single-molecule real-time sequencing and high-resolution optical mapping. Relative to the previous reference genome, our assembly features a 52-fold increase in contig length and notable improvements in the assembly of intergenic spaces and centromeres. Characterization of the repetitive portion of the genome revealed more than 130,000 intact transposable elements, allowing us to identify transposable element lineage expansions that are unique to maize. Gene annotations were updated using 111,000 full-length transcripts obtained by single-molecule real-time sequencing. In addition, comparative optical mapping of two other inbred maize lines revealed a prevalence of deletions in regions of low gene density and maize lineage-specific genes.

/pdf/improved-maize-reference-genome-with-single-molecule-9lmtaai2au.pdf

Improved maize reference genome with single-molecule technologies

Maize kernel oil is a valuable source of nutrition. Here we extensively examine the genetic architecture of maize oil biosynthesis in a genome-wide association study using 1.03 million SNPs characterized in 368 maize inbred lines, including ‘high-oil’ lines. We identified 74 loci significantly associated with kernel oil concentration and fatty acid composition (P < 1.8 × 10 −6 ), which we subsequently examined using expression quantitative trait loci (QTL) mapping, linkage mapping and coexpression analysis. More than half of the identified loci localized in mapped QTL intervals, and one-third of the candidate genes were annotated as enzymes in the oil metabolic pathway. The 26 loci associated with oil concentration could explain up to 83% of the phenotypic variation using a simple additive model. Our results provide insights into the genetic basis of oil biosynthesis in maize kernels and may facilitate marker-based breeding for oil quantity and quality.

/pdf/genome-wide-association-study-dissects-the-genetic-1g1h5gsuq7.pdf

Genome-wide association study dissects the genetic architecture of oil biosynthesis in maize kernels

The allohexaploid bread wheat genome consists of three closely related subgenomes (A, B, and D), but a clear understanding of their phylogenetic history has been lacking. We used genome assemblies of bread wheat and five diploid relatives to analyze genome-wide samples of gene trees, as well as to estimate evolutionary relatedness and divergence times. We show that the A and B genomes diverged from a common ancestor ~7 million years ago and that these genomes gave rise to the D genome through homoploid hybrid speciation 1 to 2 million years later. Our findings imply that the present-day bread wheat genome is a product of multiple rounds of hybrid speciation (homoploid and polyploid) and lay the foundation for a new framework for understanding the wheat genome as a multilevel phylogenetic mosaic.

https://science.sciencemag.org/content/sci/345/6194/1250092.full.pdf

Ancient hybridizations among the ancestral genomes of bread wheat

SNP genotyping arrays have been useful for many applications that require a large number of molecular markers such as high-density genetic mapping, genome-wide association studies (GWAS), and genomic selection. We report the establishment of a large maize SNP array and its use for diversity analysis and high density linkage mapping. The markers, taken from more than 800,000 SNPs, were selected to be preferentially located in genes and evenly distributed across the genome. The array was tested with a set of maize germplasm including North American and European inbred lines, parent/F1 combinations, and distantly related teosinte material. A total of 49,585 markers, including 33,417 within 17,520 different genes and 16,168 outside genes, were of good quality for genotyping, with an average failure rate of 4% and rates up to 8% in specific germplasm. To demonstrate this array's use in genetic mapping and for the independent validation of the B73 sequence assembly, two intermated maize recombinant inbred line populations – IBM (B73×Mo17) and LHRF (F2×F252) – were genotyped to establish two high density linkage maps with 20,913 and 14,524 markers respectively. 172 mapped markers were absent in the current B73 assembly and their placement can be used for future improvements of the B73 reference sequence. Colinearity of the genetic and physical maps was mostly conserved with some exceptions that suggest errors in the B73 assembly. Five major regions containing non-colinearities were identified on chromosomes 2, 3, 6, 7 and 9, and are supported by both independent genetic maps. Four additional non-colinear regions were found on the LHRF map only; they may be due to a lower density of IBM markers in those regions or to true structural rearrangements between lines. Given the array's high quality, it will be a valuable resource for maize genetics and many aspects of maize breeding.

http://www.alliot.fr/BIO/PDF/journal.pone.0028334.pdf

A large maize (Zea mays L.) SNP genotyping array: development and germplasm genotyping, and genetic mapping to compare with the B73 reference genome.

Genotyping with large numbers of molecular markers is now an indispensable tool within plant genetics and breeding. Especially through the identification of large numbers of single nucleotide polymorphism (SNP) markers using the novel high-throughput sequencing technologies, it is now possible to reliably identify many thousands of SNPs at many different loci in a given plant genome. For a number of important crop plants, SNP markers are now being used to design genotyping arrays containing thousands of markers spread over the entire genome and to analyse large numbers of samples. In this article, we discuss aspects that should be considered during the design of such large genotyping arrays and the analysis of individuals. The fact that crop plants are also often autopolyploid or allopolyploid is given due consideration. Furthermore, we outline some potential applications of large genotyping arrays including high-density genetic mapping, characterization (fingerprinting) of genetic material and breeding-related aspects such as association studies and genomic selection.

Large SNP arrays for genotyping in crop plants

Using high-throughput DNA sequencing technologies, it is now possible to quickly and reliably identify many thousands to millions of SNPs in a species. They can subsequently serve as markers for the development of large genotyping arrays. Large numbers of individuals derived from gene banks, landraces, breeding material and varieties can be genotyped with such arrays at an extremely high marker density in a fast, efficient and highly reproducible way. Based on our experience, we provide in this chapter an overview on various aspects that have to be considered within the process of developing such genotyping arrays, including the SNP discovery and/or collection, possible selection criteria for SNPs to be put on the array, SNP scoring and allele calling as well as data assembly for the analysis of millions of genotypes. To make the best use of these genotyping data, it will be very important to establish databases containing marker data from many genotyping experiments in order to simplify downstream data processing for scientific as for breeding purposes.

High-throughput SNP Profiling of Genetic Resources in Crop Plants Using Genotyping Arrays

An introgression breeding programme was carried out to transfer the virus resistance gene AV-1pro from the wild species Asparagus prostratus to the garden asparagus Asparagus officinalis. Serious crossing barriers caused by genetic distance and different ploidy levels of the crossing parents have been overcome using embryo rescue for the F1, BC1, and BC2 generations. The male and female fertility was widely restored in BC2 and was shown to be comparable to the cultivated asparagus. Five AV-1 resistant diploid (2n = 2x = 20) BC2 plants were selected and reciprocally backcrossed with asparagus cultivars. Segregation analyses of fourteen seedborne BC3 progenies suggested a monogenic dominant inheritance of the AV-1 resistance. Genotyping by sequencing analysis gave a strong hint for location of the resistance gene on asparagus Chromosome 2. Using an Axiom single nucleotide polymorphism (SNP) genotyping array for the analysis of three BC3 families with 10 AV-1 resistant and 10 AV-1 susceptible plants each, as well as 25 asparagus cultivars, the AV-1pro locus on Chromosome 2 was further narrowed down. The SNP with the highest LOD score was converted to a kompetitive allele specific PCR (KASP) marker, shown to be useful for the further backcross programme and serving as the starting point for the development of a diagnostic marker.

https://www.frontiersin.org/articles/10.3389/fpls.2021.809069/pdf

Eva-Maria Graner

Papers

A large maize (Zea mays L.) SNP genotyping array: development and germplasm genotyping, and genetic mapping to compare with the B73 reference genome.

Large SNP arrays for genotyping in crop plants

High-throughput SNP Profiling of Genetic Resources in Crop Plants Using Genotyping Arrays

Transfer of the Dominant Virus Resistance Gene AV-1pro From Asparagus prostratus to Chromosome 2 of Garden Asparagus A. officinalis L.