# Comparing Minimally Lethal Sampling Methods for Genomics in the Eastern Oyster (Crassostrea virginica)

## TL;DR

The study finds the best way to collect DNA from oysters without killing them, helping future research on their genetics and survival.

## Contribution

Identifies mantle biopsy from relaxed oysters preserved in ethanol as the optimal non-lethal genotyping method for Eastern oysters.

## Key findings

- Mantle biopsies from relaxed oysters preserved in ethanol provided high DNA yield and low mortality.
- Oysters over 61 mm in length had over 90% survival rate with this method.
- This technique enables repeated genomic studies without significant mortality.

## Abstract

Understanding how selection acts on individual genotypes often requires genotyping at different life stages and tracking their survival under experimental conditions. However, in mollusks, such as the Eastern oyster (
Crassostrea virginica
), collecting sufficient high‐quality DNA without causing mortality remains challenging. While prior studies have explored different noninvasive genotyping methods, an optimal method hasn't been identified. Here, we tested different techniques by combining two minimally invasive accession techniques (relaxation, shell notching), four types of cell sampling (swab, mantle biopsy, hemolymph, and extrapallial fluid), and three preservation techniques (flash freezing, ethanol, and FTA card) using nine different treatment groups. We monitored mortality for 11 days after cell sampling and quantified the effects of each treatment on mortality rates, DNA quality, and DNA quantity. Our results show that mantle biopsies from relaxed oysters preserved in ethanol yielded DNA quantities comparable to the control groups and significantly higher than either of the liquid samples, while maintaining low post‐sampling mortality. Logistic model regression demonstrated that oysters from this treatment that are longer than 61 mm have a greater than 90% chance of survival. These findings establish a viable method for genotyping juvenile oysters while minimizing post‐sampling mortality, and will facilitate future studies on genomic selection, individual‐level survival responses to stress and disease, and repeated measures of gene expression.

Non‐lethal genotyping of oysters is challenging due to the invasive nature of traditional tissue sampling methods. We compared four different sampling techniques on juvenile Eastern oysters, monitoring survival and DNA quality and quantity to identify the optimal approach. Mantle tissue sampling from relaxed oysters yielded the best balance of low mortality and high DNA yield, enabling future genomic selection studies and tracking individual stress responses without causing mortality.