Blanco-Berdugo, L. | bioRxiv Science

bioRxiv2025

Increased rate of de novo single nucleotide mutation in house mice born through assisted reproduction

Approximately 2.6% of live births in the United States are conceived using assisted reproductive technologies (ARTs). While some ART procedures, including in vitro fertilization (IVF) and intracytoplasmic sperm injection, are known to alter the epigenetic landscape of early embryonic development, their impact on DNA sequence stability is unclear. Here, we leverage the strengths of the laboratory mouse model system to investigate whether a standard ART regimen--ovarian hyperstimulation, gamete isolation, IVF, embryo culture, and embryo transfer--affects genome stability. Age-matched cohorts of ART-derived and naturally conceived C57BL/6J inbred mice were reared in a controlled setting and whole genome sequenced to [~]50x coverage. Using a rigorous pipeline for de novo single nucleotide variant (dnSNV) discovery, we observe a [~]30% increase in the dnSNV rate in ART-compared to naturally-conceived mice. Analysis of the dnSNV mutation spectrum identified signature contributions related to germline DNA repair activity, affirming expectations and evidencing the quality of our dnSNV calls. We observed no enrichment of dnSNVs in specific genomic contexts, suggesting that the observed rate increase in ART-derived mice is a general genome-wide phenomenon. Similarly, we show that the developmental timing of dnSNVs is similar in ART- and natural-born cohorts. Together, our findings show that ART is moderately mutagenic in house mice and motivate future work to define the precise procedure(s) associated with this increased mutational vulnerability. While we caution that our findings cannot be immediately translated to humans, they nonetheless emphasize a pressing need for investigations on the potential mutagenicity of ART in our species. SIGNIFICANCE STATEMENTThis study investigates whether assisted reproductive technologies (ARTs) increase the risk of inherited genetic mutations in offspring. Using a well-controlled mouse model system, we compared the de novo mutation burden in genomes of mice conceived through ART to a naturally conceived cohort. We find a [~]30% increase in new DNA mutations in ART-conceived mice, suggesting that ART procedures have a genome destabilizing effect. This increase in mutation rate appears to be uniform across the genome, rather than attributable to specific genomic contexts. While we caution against the direct translation of our findings to humans, our work nonetheless highlights the need for further research into the genetic safety of ART in people.

genomics↗

bioRxiv2025

Benchmarking Genomic Variant Calling Tools in Inbred Mouse Strains: Recommendations and Considerations

With the growing affordability of whole genome sequencing, variant identification has become an increasingly common task, but there are many challenges due to both technical and biological factors. In recent years, the number of software packages available for variant calling has rapidly increased. Understanding the benefits and drawbacks of different tools is important in setting leading practices and highlighting limitations. These considerations are crucial in model organism research, as many variant calling programs assume outbred genomes and implicit heterozygosity, which may not apply to inbred laboratory models. Here, we present an analysis of variant calling tools and their performance in the simulated genomes of the C57BL/6J inbred laboratory mouse and nine non-reference laboratory strains. Our findings reveal a tradeoff between the recall and precision of tools. Balancing these considerations, we show that an optimal call set is obtained by using an ensemble approach, but specific variant calling recommendations vary by strain and analytical goals. Further, we highlight filters improving the performance of different variant calling tools, both for the discovery of rare variants and in the discovery of strain polymorphisms. In summary, our work provides best practices for calling and filtering genomic variants in inbred organisms, particularly laboratory mice. Article SummaryIdentifying mutations and rare genetic variants is a central task for modern genomics. Many computational tools exist for variant detection, but their performance varies across diverse applications. Further, few variant calling tools have been benchmarked against inbred genomes, which are commonly used for research. To address this, we evaluated five variant calling tools using simulated data from ten diverse inbred mouse strains. We show that variant detection, recall, and precision vary across tools and mouse strains, and that an ensemble approach improves confidence in detected mutations. Our findings offer a set of best practices for variant calling in inbred organisms across diverse analytical applications.

bioinformatics↗