Manuel Lamothe

Enhancing genetic mapping of complex genomes through the design of highly-multiplexed SNP arrays: application to the large and unsequenced genomes of white spruce and black spruce

BackgroundTo explore the potential value of high-throughput genotyping assays in the analysis of large and complex genomes, we designed two highly multiplexed Illumina bead arrays using the GoldenGate SNP assay for gene mapping in white spruce (Picea glauca [Moench] Voss) and black spruce (Picea mariana [Mill.] B.S.P.).ResultsEach array included 768 SNPs, identified by resequencing genomic DNA from parents of each mapping population. For white spruce and black spruce, respectively, 69.2% and 77.1% of genotyped SNPs had valid GoldenGate assay scores and segregated in the mapping populations. For each of these successful SNPs, on average, valid genotyping scores were obtained for over 99% of progeny. SNP data were integrated to pre-existing ALFP, ESTP, and SSR markers to construct two individual linkage maps and a composite map for white spruce and black spruce genomes. The white spruce composite map contained 821 markers including 348 gene loci. Also, 835 markers including 328 gene loci were positioned on the black spruce composite map. In total, 215 anchor markers (mostly gene markers) were shared between the two species. Considering lineage divergence at least 10 Myr ago between the two spruces, interspecific comparison of homoeologous linkage groups revealed remarkable synteny and marker colinearity.ConclusionThe design of customized highly multiplexed Illumina SNP arrays appears as an efficient procedure to enhance the mapping of expressed genes and make linkage maps more informative and powerful in such species with poorly known genomes. This genotyping approach will open new avenues for co-localizing candidate genes and QTLs, partial genome sequencing, and comparative mapping across conifers.

Development of high-density SNP genotyping arrays for white spruce (Picea glauca) and transferability to subtropical and nordic congeners.

High‐density SNP genotyping arrays can be designed for any species given sufficient sequence information of high quality. Two high‐density SNP arrays relying on the Infinium iSelect technology (Illumina) were designed for use in the conifer white spruce (Picea glauca). One array contained 7338 segregating SNPs representative of 2814 genes of various molecular functional classes for main uses in genetic association and population genetics studies. The other one contained 9559 segregating SNPs representative of 9543 genes for main uses in population genetics, linkage mapping of the genome and genomic prediction. The SNPs assayed were discovered from various sources of gene resequencing data. SNPs predicted from high‐quality sequences derived from genomic DNA reached a genotyping success rate of 64.7%. Nonsingleton in silico SNPs (i.e. a sequence polymorphism present in at least two reads) predicted from expressed sequenced tags obtained with the Roche 454 technology and Illumina GAII analyser resulted in a similar genotyping success rate of 71.6% when the deepest alignment was used and the most favourable SNP probe per gene was selected. A variable proportion of these SNPs was shared by other nordic and subtropical spruce species from North America and Europe. The number of shared SNPs was inversely proportional to phylogenetic divergence and standing genetic variation in the recipient species, but positively related to allele frequency in P. glauca natural populations. These validated SNP resources should open up new avenues for population genetics and comparative genetic mapping at a genomic scale in spruce species.

Complex patterns of hybridization between exotic and native North American poplar species

UNLABELLED PREMISE OF THE STUDY Poplars and their hybrids are seen as important candidates for bioenergy initiatives. However, many concerns have been raised about large-scale plantations of new poplar cultivars. The deployment of such plants with novel traits brings the risk of potential spread of novel genome regions (including exotic genes, transgenes, or other heritable modifications) into natural populations of related species. The possibility of introgression is especially high in poplars because reproductive barriers between species are weak. Knowledge of the frequency of hybridization between cultivated trees and natural populations is one important step in the risk-assessment process. • METHODS We studied the rate of spontaneous hybridization from two sexually mature poplar plantations into adjacent natural populations of Populus deltoides and P. balsamifera. The two plantations, both in eastern Canada, contain many different complex hybrid clones with components from exotic species, mostly P. nigra, P. trichocarpa, and P. maximowiczii. We analyzed 12 species-specific single nucleotide polymorphisms from six different genes in 5373 offspring sampled from the natural populations. • RESULTS Contributions from all three exotics were found in the offspring, confirming low reproductive barriers among poplar species in these sections. The frequency of hybrid offspring varied among pollen donors, recipient populations, and years. • CONCLUSIONS The remarkably high rate of hybridization that was found in the smallest natural population sampled suggests that small peripheral populations carry a higher risk of introgression. These results could be used as a starting point for developing regulatory guidelines for the introduction of plants with novel traits.

Salmonid chromosome evolution as revealed by a novel method for comparing RADseq linkage maps

Whole genome duplication (WGD) can provide material for evolutionary innovation. Family Salmonidae is ideal for studying the effects of WGD as the ancestral salmonid underwent WGD relatively recently, ∼65 Ma, then rediploidized and diversified. Extensive synteny between homologous chromosome arms occurs in extant salmonids, but each species has both conserved and unique chromosome arm fusions and fissions. Assembly of large, outbred eukaryotic genomes can be difficult, but structural rearrangements within such taxa can be investigated using linkage maps. RAD sequencing provides unprecedented ability to generate high-density linkage maps for nonmodel species, but can result in low numbers of homologous markers between species due to phylogenetic distance or differences in library preparation. Here, we generate a high-density linkage map (3,826 markers) for the Salvelinus genera (Brook Charr S. fontinalis), and then identify corresponding chromosome arms among the other available salmonid high-density linkage maps, including six species of Oncorhynchus, and one species for each of Salmo, Coregonus, and the nonduplicated sister group for the salmonids, Northern Pike Esox lucius for identifying post-duplicated homeologs. To facilitate this process, we developed MapComp to identify identical and proximate (i.e. nearby) markers between linkage maps using a reference genome of a related species as an intermediate, increasing the number of comparable markers between linkage maps by 5-fold. This enabled a characterization of the most likely history of retained chromosomal rearrangements post-WGD, and several conserved chromosomal inversions. Analyses of RADseq-based linkage maps from other taxa will also benefit from MapComp, available at: https://github.com/enormandeau/mapcomp/

Novel Method for Comparing RADseq Linkage Maps Reveals Chromosome Evolution in Salmonids

Genome duplication can provide material for evolutionary innovation, and much remains unknown about its functional effects. Assembly of large, outbred eukaryotic genomes is difficult, but structural rearrangements within such taxa can be investigated using linkage maps. RAD sequencing provides unprecedented ability to generate high-density linkage maps for non-model species. However, these methods often result in a low number of homologous markers between species due to phylogenetic distance or technical differences in library preparation. Family Salmonidae is ideal for studying the effects of whole genome duplication. The ancestral salmonid underwent whole genome duplication around 65 million years ago and the tetraploid genome has undergone rediploidization during the salmonid diversification. In the salmonids, synteny occurs between orthologous chromosomes, but each species exhibits conserved and unique chromosome arm fusions and fissions. In this study, we identify orthologous chromosome arms within the salmonids using available RADseq salmonid linkage maps along with a new high-density linkage map (3826 markers) constructed for the Salvelinus genera (Brook Charr S. fontinalis ). We developed MapComp, a program that identifies identical and proximal markers between linkage maps using a reference genome of a related species as an intermediate (e.g. Rainbow Trout Oncorhynchus mykiss ). We greatly increased the number of comparable markers between linkage maps relative to that obtained using only identical markers. This enabled a characterization of the most likely history of retained chromosomal rearrangements post-whole genome duplication in five species of Oncorhynchus , and one species of each of Salvelinus , Salmo , and Coregonus , representing all of the main salmonid genera. Additionally, integration with the genetic map of the pre-duplicated sister species Northern Pike Esox lucius permitted the identification of homeologous chromosomes in all species. Putative conserved inversions within chromosome arms were also identified among species. Analyses of RADseq-based linkage maps from other taxa are likely to benefit from MapComp, available at: https://github.com/enormandeau/mapcomp/.Whole genome duplication (WGD) can provide material for evolutionary innovation. Assembly of large, outbred eukaryotic genomes can be difficult, but structural rearrangements within such taxa can be investigated using linkage maps. RAD sequencing provides unprecedented ability to generate high-density linkage maps for non-model species, but can result in low numbers of homologous markers between species due to phylogenetic distance or differences in library preparation. Family Salmonidae is ideal for studying the effects of WGD as the ancestral salmonid underwent WGD relatively recently, around 65 million years ago, then rediploidized and diversified. Extensive synteny between orthologous chromosomes occurs in extant salmonids, but each species has both conserved and unique chromosome arm fusions and fissions. Here we generate a high-density linkage map (3826 markers) for the Salvelinus genera (Brook Charr S. fontinalis), and then identify orthologous chromosome arms among the other available salmonid high-density linkage maps, including six species of Oncorhynchus, and one species for each of Salmo and Coregonus, as well as the sister group for the salmonids, Esox lucius for homeolog designation. To this end, we developed MapComp, a program that identifies identical and proximal markers between linkage maps using a reference genome of a related species as an intermediate. This approach increases the number of comparable markers between linkage maps by 5-fold, enabling a characterization of the most likely history of retained chromosomal rearrangements post-WGD, and identifying several conserved chromosomal inversions. Analyses of RADseq-based linkage maps from other taxa will also benefit from MapComp, available at: https://github.com/enormandeau/mapcomp/

Gene copy number variations in adaptive evolution: The genomic distribution of gene copy number variations revealed by genetic mapping and their adaptive role in an undomesticated species, white spruce (Picea glauca)

Gene copy number variation (CNV) has been associated with phenotypic variability in animals and plants, but a genomewide understanding of their impacts on phenotypes is largely restricted to human and agricultural systems. As such, CNVs have rarely been considered in investigations of the genomic architecture of adaptation in wild species. Here, we report on the genetic mapping of gene CNVs in white spruce, which lacks a contiguous assembly of its large genome (~20 Gb), and their relationships with adaptive phenotypic variation. We detected 3,911 gene CNVs including de novo structural variations using comparative genome hybridization on arrays (aCGH) in a large progeny set. We inferred the heterozygosity at CNV loci within parents by comparing haploid and diploid tissues and genetically mapped 82 gene CNVs. Our analysis showed that CNVs were distributed over 10 linkage groups and identified four CNV hotspots that we predict to occur in other species of the Pinaceae. Significant relationships were found between 29 of the gene CNVs and adaptive traits based on regression analyses with timings of bud set and bud flush, and height growth, suggesting a role for CNVs in climate adaptation. The importance of CNVs in adaptive evolution of white spruce was also indicated by functional gene annotations and the clustering of 31% of the mapped adaptive gene CNVs in CNV hotspots. Taken together, these results illustrate the feasibility of studying CNVs in undomesticated species and represent a major step towards a better understanding of the roles of CNVs in adaptive evolution.

History rather than hybridization determines population structure and adaptation in Populus balsamifera

Hybridization between species is known to greatly affect their genetic diversity and, therefore, their evolution. Also, within species, there may be genetic clusters between which gene flow is limited, which may impact natural selection. However, few studies have looked simultaneously at the influence of among‐species and within‐species gene flow. Here, we study the influence of hybridization between Populus balsamifera and Populus trichocarpa on population structure and adaptation in P. balsamifera. We did this by sampling a total of 1517 individuals from across the ranges of these two species, and by genotyping them using a combination of 93 nuclear and 17 cpDNA SNPs. We found that hybridization is mostly limited to the contact zone where the species’ distributions overlap. Within P. balsamifera, we found multiple levels of population structure. Interestingly, the border between the Eastern and Central clusters is very sharp, whereas the border between the Central and Western clusters is diffuse. Outlier analysis revealed that three loci associated with the sharp border were also associated with climate. We hypothesize that the observed clusters derive from three refugia during the Pleistocene ice ages. Between the Central and Western clusters, post‐glacial long‐distance gene flow has led to the diffusion of their border. In the Eastern cluster, we hypothesize that endogenous genomic barriers have developed, leading to the sharp border and a spurious climate association. We conclude that the large‐scale genetic structure of P. balsamifera is mostly shaped by historical factors and the influence of interspecific hybridization is limited.