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

Abstract

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% of the gaps in the existing reference genome assembly, improving contiguity over five-fold. 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.

Chromosomal Fusions Facilitate Adaptation to Divergent Environments in Threespine Stickleback

“…To distinguish between these possibilities, we used the newly available whole-genome assemblies of the outgroup Aul. flavidus (Li Q, Lindtke D, Rodríguez-Ramírez C, Kakioka R, Takahashi H, Toyoda A, Kitano J, Ehrlich RL, Mell JC, Yeaman S, personal communication), P. pungitius ( Varadharajan et al 2019 ), and G. aculeatus ( Nath et al 2021 ), as well as the high-quality assembly of A. quadracus generated in this study. We then focused on the whole-chromosome rearrangements that have occurred in G. aculeatus to determine whether these rearrangements are associated with genetic loci that underlie adaptation to divergent marine and freshwater habitats in this species.…”

“…The N50 length is 18.10 Mb, and the assembly quality assessed by BUSCO was relatively high with 96.9% completeness. Apeltes quadracus has a smaller genome than the other existing stickleback genome assemblies (∼449 Mb for G. aculeatus ; Nath et al [2021 ] and ∼521 Mb for P. pungitius ; Varadharajan et al [2019 ]). We constructed a repeat library for A. quadracus using de novo and homology-based approaches (see Materials and Methods).…”

Heterogeneous Histories of Recombination Suppression on Stickleback Sex Chromosomes

“…A fifth version of the assembly of the threespine stickleback genome was published at https://stickleback.genetics.uga.edu/ , whereas this manuscript was in review ( Nath et al 2021 ). Importantly, it corrected numerous annotation errors in the earlier assembly that precluded accurate analyses of the coding regions.…”

“…To quantify Y chromosome degeneration, we calculated the mean read depth ratio for each gene from data mapped to the fifth assembly of the threespine stickleback genome ( Nath et al 2021 ). Read depth for males and females was calculated using VCFtools –geno-depth v.1.15 ( Danecek et al 2011 ).…”

“…We calculated genomic divergence using a pipeline and scripts developed by Dixon et al (2019) . This analysis used sequencing reads mapped to the fifth assembly of the threespine stickleback genome and the corresponding Ensembl annotations ( Nath et al 2021 ). First, we generated consensus sequences for the blackspotted X, blackspotted Y, and threespine X sequences from our pedigrees.…”