[Paper Review] Using sequencing coverage statistics to identify sex chromosomes in minke whales

Ketil Malde, Rasmus Skern+1 more|arXiv (Cornell University)|Feb 18, 2019

Marine animal studies overviewEnvironmental Science43 references3 citations

TL;DR

This study presents a simple, coverage-based bioinformatic method to identify sex chromosomes in minke whales using publicly available whole-genome sequencing data. By analyzing sequencing depth differences between male and female samples, the authors successfully classified genomic scaffolds as X or Y chromosomes, identifying ~90 Mbp of sex-linked sequences—demonstrating that coverage statistics alone can reliably detect sex chromosomes without prior knowledge or complex homology searches.

ABSTRACT

The ever-increasing number of genome sequencing and resequencing projects is a central source of insights into the ecology and evolution of non-model organisms. An important aspect of genomics is the elucidation of sex determination systems and identifying genes on sex chromosomes. This not only helps reveal mechanisms behind sex determination in the species under study, but their characteristics make sex chromosomes a unique tool for studying the mechanisms and effects of recombination and genomic rearrangements and how they affect adaption and selection. Despite this, many sequencing projects omit such investigations. Here, we apply a simple method using sequencing coverage statistics to identify scaffolds belonging to the sex chromosomes of minke whale, and show how the sex chromosome system can be determined using coverage statistics alone. Using publicly available data, we identify the previously unknown sex of an Antarctic minke whale as female. We further investigate public sequence data from the different species and sub-species of minke whale, and classify genomic scaffolds from a published minke whale assembly as X or Y chromosomal sequences. Our findings are consistent with previous results that identified a handful of scaffolds as sex chromosomal, but we are able to identify a much larger set of scaffolds, likely to represent close to the complete sex chromosomal sequences for the minke whale. Sequence coverage statistics provides a readily available tool for investigating the sex determination system and locate genes on sex chromosomes. This analysis is straightforward and can often be performed with existing resources.

Motivation & Objective

• To develop a low-cost, accessible method for identifying sex chromosomes in non-model organisms using only sequencing coverage data.
• To resolve the sex of an Antarctic minke whale using publicly available genomic data, confirming it as female.
• To classify genomic scaffolds from a published minke whale assembly as X or Y chromosomal sequences without relying on homology-based methods.
• To demonstrate that coverage-based analysis can identify a much larger set of sex chromosome scaffolds than previous homology-based approaches.
• To provide a scalable, computationally simple approach applicable to any species with XY or ZW sex chromosome systems using existing genomic resources.

Proposed method

• Utilize whole-genome shotgun sequencing data from male and female minke whales to compute read coverage across genomic scaffolds.
• Compare coverage distributions between male and female samples to identify scaffolds with hemizygous patterns characteristic of Y (or W) chromosomes.
• Apply a clustering approach to group scaffolds by coverage level, distinguishing autosomes (balanced coverage) from sex chromosomes (asymmetric coverage).
• Use scaffold length as a covariate to correct for coverage bias, improving accuracy in coverage estimation.
• Apply statistical modeling (e.g., negative binomial or mixture models) to account for overdispersion and heteroskedasticity in coverage data.
• Validate results using publicly available data and release scripts for reproducibility via GitHub.

Experimental results

Research questions

• RQ1 Can sequencing coverage statistics alone reliably identify sex chromosomes in minke whales without prior knowledge of the sex chromosome system?
• RQ2 How many scaffolds in the minke whale genome can be confidently assigned to the X or Y chromosome using coverage-based classification?
• RQ3 To what extent does this method outperform homology-based approaches in identifying sex-linked sequences?
• RQ4 Can this method be applied to resolve the sex of previously unclassified individuals using only public sequencing data?
• RQ5 What is the estimated size and gene content of the minke whale sex chromosomes based on coverage patterns?

Key findings

• The method successfully identified 90 Mbp of genomic scaffolds as belonging to the sex chromosomes, representing a substantial increase over previous homology-based identifications.
• The previously unknown sex of an Antarctic minke whale was determined as female using only coverage statistics from public sequencing data.
• The analysis classified a large number of scaffolds as X or Y chromosomal sequences, with coverage patterns consistent across multiple species and subspecies of minke whale.
• The approach identified a much larger set of sex-linked scaffolds than earlier studies, suggesting near-complete coverage of the sex chromosome regions.
• The method is robust even with a single sequencing library per sex and works effectively on high-quality genome assemblies with long scaffolds (>10 kbp).
• The results were validated using publicly available data, and analysis scripts are openly shared for reproducibility and broader application.