Improved contiguity of the threespine stickleback genome using long-read sequencing

While the cost and time for assembling a genome has drastically decreased, it still remains a challenge to assemble a highly contiguous genome. These challenges are rapidly being overcome by the integration of long-read sequencing technologies. Here, we use long-read sequencing to improve the contiguity of the threespine stickleback fish (Gasterosteus aculeatus) genome, a prominent genetic model species. Using Pacific Biosciences sequencing, we assembled a highly contiguous genome of a freshwater fish from Paxton Lake. Using contigs from this genome, we were able to fill over 76.7% of the gaps in the existing reference genome assembly, improving contiguity over fivefold. Our gap filling approach was highly accurate, validated by 10X Genomics long-distance linked-reads. In addition to closing a majority of gaps, we were able to assemble segments of telomeres and centromeres throughout the genome. This highlights the power of using long sequencing reads to assemble highly repetitive and difficult to assemble regions of genomes. This latest genome build has been released through a newly designed community genome browser that aims to consolidate the growing number of genomics datasets available for the threespine stickleback fish.

Improved contiguity of the threespine stickleback genome using long-read sequencing

While the cost and time for assembling a genome have drastically reduced, it still remains a challenge to assemble a highly contiguous genome. These challenges are rapidly being overcome by the integration of long-read sequencing technologies. Here, we use long sequencing reads to improve the contiguity of the threespine stickleback fish (Gasterosteus aculeatus) genome, a prominent genetic model species. Using Pacific Biosciences sequencing, we were able to fill over 76% of the gaps in the genome, improving contiguity over five-fold. Our approach was highly accurate, validated by 10X Genomics long-distance linked-reads. In addition to closing a majority of gaps, we were able to assemble segments of telomeres and centromeres throughout the genome. This highlights the power of using long sequencing reads to assemble highly repetitive and difficult to assemble regions of genomes. This latest genome build has been released through a newly designed community genome browser that aims to consolidate the growing number of genomics datasets available for the threespine stickleback fish.

Selection, linkage, and population structure interact to shape genetic variation among threespine stickleback genomes

The outcome of selection on genetic variation depends on the geographic organization of individuals and populations as well as the syntenic organization of loci within the genome. Spatially variable selection between marine and freshwater habitats has had a significant and heterogeneous impact on patterns of genetic variation across the genome of threespine stickleback fish. When marine stickleback invade freshwater habitats, more than a quarter of the genome can respond to divergent selection, even in as little as 50 years. This process largely uses standing genetic variation that can be found ubiquitously at low frequency in marine populations, can be millions of years old, and is likely maintained by significant bidirectional gene flow. Here, we combine population genomic data of marine and freshwater stickleback from Cook Inlet, Alaska, with genetic maps of stickleback fish derived from those same populations to examine how linkage to loci under selection affects genetic variation across the stickleback genome. Divergent selection has had opposing effects on linked genetic variation on chromosomes from marine and freshwater stickleback populations: near loci under selection, marine chromosomes are depauperate of variation while these same regions among freshwater genomes are the most genetically diverse. Forward genetic simulations recapitulate this pattern when different selective environments also differ in population structure. Lastly, dense genetic maps demonstrate that the interaction between selection and population structure may impact large stretches of the stickleback genome. These findings advance our understanding of how the structuring of populations across geography influences the outcomes of selection, and how the recombination landscape broadens the genomic reach of selection.

Ancient genomic variation underlies repeated ecological adaptation in young stickleback populations

ABSTRACTAdaptation in the wild often involves standing genetic variation (SGV), which allows rapid responses to selection on ecological timescales. However, we still know little about how the evolutionary histories and genomic distributions of SGV influence local adaptation in natural populations. Here, we address this knowledge gap using the threespine stickleback fish (Gasterosteus aculeatus) as a model. We extend the popular restriction site-associated DNA sequencing (RAD-seq) method to produce phased haplotypes approaching 700 base pairs (bp) in length at each of over 50,000 loci across the stickleback genome. Parallel adaptation in two geographically isolated freshwater pond populations consistently involved fixation of haplotypes that are identical-by-descent. In these same genomic regions, sequence divergence between marine and freshwater stickleback, as measured by dXY, reaches ten-fold higher than background levels and structures genomic variation into distinct marine and freshwater haplogroups. By combining this dataset with a de novo genome assembly of a related species, the ninespine stickleback (Pungitius pungitius), we find that this habitat-associated divergent variation averages six million years old, nearly twice the genome-wide average. The genomic variation that is involved in recent and rapid local adaptation in stickleback has actually been evolving throughout the 15-million-year history since the two species lineages split. This long history of genomic divergence has maintained large genomic regions of ancient ancestry that include multiple chromosomal inversions and extensive linked variation. These discoveries of ancient genetic variation spread broadly across the genome in stickleback demonstrate how selection on ecological timescales is a result of genome evolution over geological timescales, and vice versa.IMPACT STATEMENTAdaptation to changing environments requires a source of genetic variation. When environments change quickly, species often rely on variation that is already present – so-called standing genetic variation – because new adaptive mutations are just too rare. The threespine stickleback, a small fish species living throughout the Northern Hemisphere, is well-known for its ability to rapidly adapt to new environments. Populations living in coastal oceans are heavily armored with bony plates and spines that protect them from predators. These marine populations have repeatedly invaded and adapted to freshwater environments, losing much of their armor and changing in shape, size, color, and behavior.Adaptation to freshwater environments can occur in mere decades and probably involves lots of standing genetic variation. Indeed, one of the clearest examples we have of adaptation from standing genetic variation comes from a gene, eda, that controls the shifts in armor plating. This discovery involved two surprises that continue to shape our understanding of the genetics of adaptation. First, freshwater stickleback from across the Northern Hemisphere share the same version, or allele, of this gene. Second, the ‘marine’ and ‘freshwater’ alleles arose millions of years ago, even though the freshwater populations studied arose much more recently. While it has been hypothesized that other genes in the stickleback genome may share these patterns, large-scale surveys of genomic variation have been unable to test this prediction directly.Here, we use new sequencing technologies to survey DNA sequence variation across the stickleback genome for patterns like those at the eda gene. We find that every region of the genome associated with marine-freshwater genetic differences shares this pattern to some degree. Moreover, many of these regions are as old or older than eda, stretching back over 10 million years in the past and perhaps even predating the species we now call the threespine stickleback. We conclude that natural selection has maintained this variation over geological timescales and that the same alleles we observe in freshwater stickleback today are the same as those under selection in ancient, now-extinct freshwater habitats. Our findings highlight the need to understand evolution on macroevolutionary timescales to understand and predict adaptation happening in the present day.

Co-localization of growth QTL with differentially expressed candidate genes in rainbow trout

We tested whether genes differentially expressed between large and small rainbow trout co-localized with familial QTL regions for body size. Eleven chromosomes, known from previous work to house QTL for weight and length in rainbow trout, were examined for QTL in half-sibling families produced in September (1 XY male and 1 XX neomale) and December (1 XY male). In previous studies, we identified 108 candidate genes for growth expressed in the liver and white muscle in a subset of the fish used in this study. These gene sequences were BLASTN aligned against the rainbow trout and stickleback genomes to determine their location (rainbow trout) and inferred location based on synteny with the stickleback genome. Across the progeny of all three males used in the study, 63.9% of the genes with differential expression appear to co-localize with the QTL regions on 6 of the 11 chromosomes tested in these males. Genes that co-localized with QTL in the mixed-sex offspring of the two XY males primarily showed up-regulation in the muscle of large fish and were related to muscle growth, metabolism, and the stress response.