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Adaptive evolution often involves structural variation affecting genes or cis-regulatory changes that engender novel and favorable gain-of-function gene regulation. Such mutation could result in a favorable dominant trait. At the same time, the gene product could be dosage sensitive if its change in concentration disrupts another trait. As a result, the mutant allele would display dosage-sensitive pleiotropy (DSP). By minimizing imbalance while conserving the favorable dominant effect, heterozygosity can increase fitness and result in heterosis. The properties of these alleles are consistent with evidence from multiple studies that indicate increased fitness of heterozygous regulatory mutations. DSP can help explain mysterious properties of heterosis as well as other effects of hybridization. 

Ancient whole-genome duplications (WGDs) are believed to facilitate novelty and adaptation by providing the raw fuel for new genes. However, it is unclear how recent WGDs may contribute to evolvability within recent polyploids. Hybridization accompanying some WGDs may combine divergent gene content among diploid species. Some theory and evidence suggest that polyploids have a greater accumulation and tolerance of gene presence-absence and genomic structural variation, but it is unclear to what extent either is true. To test how recent polyploidy may influence pangenomic variation, we sequenced, assembled, and annotated twelve complete, chromosome-scale genomes of Camelina sativa, an allohexaploid biofuel crop with three distinct subgenomes. Using pangenomic comparative analyses, we characterized gene presence-absence and genomic structural variation both within and between the subgenomes. We found over 75% of ortholog gene clusters are core in Camelina sativa and <10% of sequence space was affected by genomic structural rearrangements. In contrast, 19% of gene clusters were unique to one subgenome, and the majority of these were Camelina-specific (no ortholog in Arabidopsis). We identified an inversion that may contribute to vernalization requirements in winter-type Camelina, and an enrichment of Camelina-specific genes with enzymatic processes related to seed oil quality and Camelina’s unique glucosinolate profile. Genes related to these traits exhibited little presence-absence variation. Our results reveal minimal pangenomic variation in this species, and instead show how hybridization accompanied by WGD may benefit polyploids by merging diverged gene content of different species.

 

Conversion of potato from a tetraploid, heterozygous, vegetatively propagated crop to a diploid F1 hybrid, propagated via botanical seed, would constitute a considerable advance for global agriculture, but faces multiple challenges. One such challenge is the difficulty in inbreeding potato, which involves purging deleterious alleles from its genome. This commentary discusses possible reasons for this difficulty and highlights a recent sequence-based effort to classify SNP variation, in potato germplasm, according to its deleterious potential. Tools and strategies connected to this database may facilitate development of F1 hybrids.

 

High-throughput sequencing-based methods for bulked segregant analysis (BSA) allow for the rapid identification of genetic markers associated with traits of interest. BSA studies have successfully identified qualitative (binary) and quantitative trait loci (QTLs) using QTL mapping. However, most require population structures that fit the models available and a reference genome. Instead, high-throughput short-read sequencing can be combined with BSA of k-mers (BSA-k-mer) to map traits that appear refractory to standard approaches. This method can be applied to any organism and is particularly useful for species with genomes diverged from the closest sequenced genome. It is also instrumental when dealing with highly heterozygous and potentially polyploid genomes without phased haplotype assemblies and for which a single haplotype can control a trait. Finally, it is flexible in terms of population structure. Here, we apply the BSA-k-mer method for the rapid identification of candidate regions related to seed spot and seed size in diploid potato. Using a mixture of F1 and F2 individuals from a cross between 2 highly heterozygous parents, candidate sequences were identified for each trait using the BSA-k-mer approach. Using parental reads, we were able to determine the parental origin of the loci. Finally, we mapped the identified k-mers to a closely related potato genome to validate the method and determine the genomic loci underlying these sequences. The location identified for the seed spot matches with previously identified loci associated with pigmentation in potato. The loci associated with seed size are novel. Both loci are relevant in future breeding toward true seeds in potato.

 

Abstract Large-scale structural variations, such as chromosomal translocations, can have profound effects on fitness and phenotype, but are difficult to identify and characterize. Here, we describe a simple and effective method aimed at identifying translocations using only the dosage of sequence reads mapped on the reference genome. We binned reads on genomic segments sized according to sequencing coverage and identified instances when copy number segregated in populations. For each dosage-polymorphic 1 Mb bin, we tested independence, effectively an apparent linkage disequilibrium (LD), with other variable bins. In nine potato (Solanum tuberosum) dihaploid families translocations affecting pericentromeric regions were common and in two cases were due to genomic misassembly. In two populations, we found evidence for translocation affecting euchromatic arms. In cv. PI 310467, a nonreciprocal translocation between chromosomes (chr.) 7 and 8 resulted in a 5–3 copy number change affecting several Mb at the respective chromosome tips. In cv. “Alca Tarma,” the terminal arm of chr. 4 translocated to the tip of chr. 1. Using oligonucleotide-based fluorescent in situ hybridization painting probes (oligo-FISH), we tested and confirmed the predicted arrangement in PI 310467. In 192 natural accessions of Arabidopsis thaliana, dosage haplotypes tended to vary continuously and resulted in higher noise, while apparent LD between pericentromeric regions suggested the effect of repeats. This method, LD-CNV, should be useful in species where translocations are suspected because it tests linkage without the need for genotyping. 

Dihaploid production from elite tetraploid cultivars is key to both traditional and novel breeding approaches that seek to simplify potato genetics. For this purpose, efficient and widely compatible haploid inducers (HIs) are needed. We compared PL-4, a new HI developed at the International Potato Center, to known HIs IvP101 and IvP35. By pollination of elite tetraploid breeding lines, we showed that PL-4 performed significantly better and had a homogeneous response regardless of the genetic background of the pistillate parents, on the most important efficiency traits—number of dihaploids per 100 fruits and haploid induction rate. Moreover, PL-4 exhibited a reduced proportion of hybrid seeds, a convenient trait for efficient screening. In this context, we recommend PL-4 as an enhanced HI for the potato breeding community.

 

The Targeting Induced Local Lesions in Genomes (TILLING) technology is a reverse genetic strategy broadly applicable to every kind of genome and represents an attractive tool for functional genomic and agronomic applications. It consists of chemical random mutagenesis followed by high-throughput screening of point mutations in targeted genomic regions. Although multiple methods for mutation discovery in amplicons have been described, next-generation sequencing (NGS) is the tool of choice for mutation detection because it quickly allows for the analysis of a large number of amplicons. The aim of the present work was to screen a previously generated sunflower TILLING population and identify alterations in genes involved in several important and complex physiological processes. Twenty-one candidate sunflower genes were chosen as targets for the screening. The TILLING by sequencing strategy allowed us to identify multiple mutations in selected genes and we subsequently validated 16 mutations in 11 different genes through Sanger sequencing. In addition to addressing challenges posed by outcrossing, our detection and validation of mutations in multiple regulatory loci highlights the importance of this sunflower population as a genetic resource.

 

Abstract In cultivated tetraploid potato (Solanum tuberosum), reduction to diploidy (dihaploidy) allows for hybridization to diploids and introgression breeding and may facilitate the production of inbreds. Pollination with haploid inducers yields maternal dihaploids, as well as triploid and tetraploid hybrids. Dihaploids may result from parthenogenesis, entailing the development of embryos from unfertilized eggs, or genome elimination, entailing missegregation and the loss of paternal chromosomes. A sign of genome elimination is the occasional persistence of haploid inducer DNA in some dihaploids. We characterized the genomes of 919 putative dihaploids and 134 hybrids produced by pollinating tetraploid clones with three haploid inducers: IVP35, IVP101, and PL-4. Whole-chromosome or segmental aneuploidy was observed in 76 dihaploids, with karyotypes ranging from 2n=2x-1=23 to 2n=2x+3=27. Of the additional chromosomes in 74 aneuploids, 66 were from the non-inducer parent and 8 from the inducer parent. Overall, we detected full or partial chromosomes from the haploid inducer parent in 0.87% of the dihaploids, irrespective of parental genotypes. Chromosomal breaks commonly affected the paternal genome in the dihaploid and tetraploid progeny, but not in the triploid progeny, correlating instability to sperm ploidy and to haploid induction. The residual haploid inducer DNA discovered in the progeny is consistent with genome elimination as the mechanism of haploid induction.