Dataset: Oyster genetic assignment data from an experimental oyster reef restoration in Ninigret Pond, Rhode Island (USA) in 2017-2020

These data were published in Truskey et al. 2025 (Evolutionary Applications). All figure numbers and supplementary materials mentioned refer to Truskey et al. 2025 (Evolutionary Applications).

Restoration experiment and sample collection

In April 2017, we collaborated with local oyster farmers to source oyster eyed-larvae from four commercial hatcheries for the construction of new experimental oyster reefs in Ninigret Pond (Charlestown, Rhode Island, USA). Hatchery sources were selected across a broad geographic sampling range to maximize diversity (Figure 1a of Truskey et al. 2025). They included two regional hatcheries, one from Massachusetts (MA) and one from New York (NY), and two more distant hatcheries, one from Maine (ME) and one from Virginia (VA). A local Rhode Island hatchery facility received oyster eyed-larvae from each source hatchery, set the eyed-larvae on dead oyster and/or clam shell (hereafter referred to as spat-on-shell oysters), and distributed the spat-on-shell oysters to three local oyster growers (Grower 1: MA; Grower 2: ME, NY, VA; Grower 3: NY, VA) who maintained these juvenile oysters on separate leased oyster farm plots prior to reef construction. Immediately prior to reef construction, we collected 20 juvenile spat-on-shell oysters from each grower-hatchery source combination for genetic analysis (n = 6 combinations; Grower 1-MA; Grower 2-ME; Grower 2-NY; Grower 2-VA; Grower 3-NY; Grower 3-VA).

In October and November 2017, we constructed 16 subtidal reefs within a no-harvest Shellfish Management Area in Ninigret Pond. Reefs were created across four 0.025-acre experimental blocks, with four reefs per block (Figure 2 of Truskey et al. 2025). Each reef consisted of a base layer of 0.25 cubic yards of dead oyster or clam shell deployed in October and topped with 1.25 cubic yards of spat-on-shell oysters in early November from the stock grown out by the local oyster growers. Our experiment was originally designed to seed three reefs per block with a different single hatchery source from the four available hatchery sources and to seed the fourth reef per block with a mixed combination of the three sources (e.g., Reef 1: ME, Reef 2: MA, Reef 3: NY, Reef 4: ME+MA+NY). However, analysis of the hatchery samples from oyster growers collected prior to reef construction revealed an early, unintended mixing of some of the sources (Appendix S1 of Truskey et al. 2025; Figure S1 of Truskey et al. 2025), resulting in mixtures of multiple sources on 12 out of the 16 constructed reefs. Thus, we focused our analyses on the actual genetic composition of each reef as determined by genetic sampling at two time points (fall 2018, fall 2020). 

In fall 2018, we haphazardly sampled live oysters from each reef on scuba or snorkel (N = 512 individuals total, 32 per reef). Oysters were put on ice and transported to the Northeastern University Marine Science Center where they were held at −80°C until DNA extraction. By the fall of 2020, live oyster densities on all experimental reefs had declined, reflecting mortality of the original planted oysters and a lack of recruitment, consistent with other data from this system (Barrett et al. 2024). To assess whether this mortality was associated with a consistent change in the genetic composition of surviving oysters on reefs, we repeated our sampling in fall 2020 and compared the resulting reef genetic profiles to those from fall 2018. Reef sample sizes varied at this time point due to low live abundances (N = 249 individuals total, ranging from 8 to 32 per reef).

DNA extraction, RADseq library preparation, and bioinformatics

Genomic DNA was extracted using the E-Z 96 Tissue DNA Kit (Omega-Biotek, Norcross, GA) following the animal tissue protocol with tissue centrifugation. Double-digest restriction-site-associated DNA (ddRAD) libraries with individually barcoded samples were prepared in three batches following Parchman et al. (2012): (1) initial hatchery samples (n = 120), (2) fall 2018 reef samples (n = 480; 30 oysters per reef across 16 reefs), and (3) fall 2020 reef samples (n = 248). For additional details on ddRAD library preparation, see Appendix S2 of Truskey et al. 2025. All libraries were sequenced with 100-bp single-end reads on an Illumina platform. The initial and fall 2020 batches were sequenced on a single lane of the Illumina HiSeq 2500 at Tufts University Core Facility Genomics; the fall 2018 batch was sequenced on two lanes of the Illumina NovaSeq 6000 at the University of Texas at Austin Genomic Sequencing and Analysis Facility.