A second unveiling: Haplotig masking of the eastern oyster genome improves population‐level inference

Abstract

Genome assembly can be challenging for species that are characterized by high amounts of polymorphism, heterozygosity, and large effective population sizes. High levels of heterozygosity can result in genome mis‐assemblies and a larger than expected genome size due to the haplotig versions of a single locus being assembled as separate loci. Here, we describe the first chromosome‐level genome for the eastern oyster, Crassostrea virginica. Publicly released and annotated in 2017, the assembly has a scaffold N50 of 54 mb and is over 97.3% complete based on BUSCO analysis. The genome assembly for the eastern oyster is a critical resource for foundational research into molluscan adaptation to a changing environment and for selective breeding for the aquaculture industry. Subsequent resequencing data suggested the presence of haplotigs in the original assembly, and we developed a post hoc method to break up chimeric contigs and mask haplotigs in published heterozygous genomes and evaluated improvements to the accuracy of downstream analysis. Masking haplotigs had a large impact on SNP discovery and estimates of nucleotide diversity and had more subtle and nuanced effects on estimates of heterozygosity, population structure analysis, and outlier detection. We show that haplotig masking can be a powerful tool for improving genomic inference, and we present an open, reproducible resource for the masking of haplotigs in any published genome.

Sex‐specific gene expression in eastern oyster, Crassostrea virginica, gonad and mantle tissues

“…By conditioning adult eastern oysters in a controlled laboratory environment and comparing female and male global expression profiles in reproductively ripe gonad and mantle tissues, we confirmed that many conserved sex differentiation genes are present and predictably expressed in eastern oysters. Deep sequencing of individual transcriptomes and subsequent mapping to the highly contiguous, chromosome-level eastern oyster genome assembly (Puritz et al, 2023) also provided opportunity for more in-depth exploration of sexspecific processes in gonad and mantle tissues. Many transcripts associated with flagellar formation, spermiogenesis, and degradation of damaged sperm were identified in male gonad, whereas those involved in cell cycle regulation were observed in female gonad.…”

“…Trimmed reads were mapped to a haplotig masked version (Puritz et al, 2023) of the Crassostrea virginica genome v3.0 (NCBI; GCA_002022765.4) using CLC Genomic Workbench's RNA-Seq Analysis tool. To account for high polymorphism in oysters (L. Zhang, Li, et al, 2014), the length fraction setting was reduced to 0.6 as in Proestou and Sullivan (2020), while all other settings were kept at default.…”

“…Here, we leveraged a high-quality annotated genome (Puritz et al, 2023), affordable Illumina-based transcriptome sequencing technology, and optimized RNAseq analysis pipelines to generate global gene expression profiles for mature eastern oyster gonad tissues. Expression profiles were used to highlight sex-specific patterns and identify conserved and novel transcripts involved in sex differentiation in the eastern oyster.…”

Consequences of domestication in eastern oyster: Insights from whole genomic analyses

“…Mitochondrial SNPs on the array were excluded. SNPs located in putative inversions (Puritz et al., 2022 , 2023 ) were excluded to prevent artifacts in population structure patterns from long‐range linkage disequilibrium. Additional filtering steps were applied using VCFtools (Danecek et al., 2011 ): excluded SNPs with minor allele frequency < 0.05 across 18 wild and domestic populations combined; individuals were removed if they had >10% missing data; SNPs with a call rate less than 95% in any single population were excluded.…”

Parallel evolution in Crassostrea oysters along the latitudinal gradient is associated with variation in multiple genes involved in adipogenesis

“…Then the phylogenetic tree was estimated using RaxML v.8.2.12 (Stamatakis, 2014). Finally, the divergence time was estimated using MCMCTree from PAML v.4.9 (Yang, 2007) (Puritz et al, 2023). The calibration divergence of C. virginica and C. angulata was estimated at 73 Ma (Plazzi & Passamonti, 2010;Ren et al, 2010).…”