Non-invasive DNA collection for parentage analysis for bivalves: A case study from the silver-lipped pearl oyster (Pinctada maxima)

Abstract

Due to the high revenues generated by the pearl industry, interest in developing breeding programs is increasing. The traditional approach for collecting DNA for pedigree analyses in breeding programs for the silver-lipped pearl oyster (Pinctada maxima) involves tissue excision from the muscular foot and/or mantle. Tissue excision causes injury and stress to the animal, and can lead to reduced growth efficiency, bacterial infection and subsequent mortality when oysters are returned to the non-sterile marine environment. The approach of swabbing tissue to collect cells for DNA analyses offers a potential non-invasive and rapid method but has not been reported previously for pearl oysters; particularly where DNA is destined for a high-throughput genotype-by-sequencing (GBS) single nucleotide polymorphism (SNP) genotyping platform. To evaluate the potential of swabbing as a method to collect DNA for downstream GBS SNP genotyping under commercial farming conditions, 29 broodstock pearl oysters were swabbed with two different swab brushes (a stiff and soft brush). Concentration of the extracted DNA from swabs was found to meet requirements for genotyping (44 to 59 ng/μL) and 260/280 ratios were in the acceptable range for DNA purity (1.9–2.2). SNP genotype calls derived from swab samples were highly correlated with those derived from tissue excised from the same oyster (mean of 0.97 ± 0.04 for soft bristle – mean of 0.98 ± 0.02 for stiff bristle). Finally, we evaluated the performance of swabbing under a commercial breeding program scenario, whereby 923 spat were genotyped and 18 months later 1927 pearl oysters from the same commercial cohort were swabbed to determine parental contributions for the cohort at the two time periods. Results demonstrated that the swab method is reliable for collecting sufficient quantity and quality of DNA for genome-wide SNP genotyping enabling accurate pedigree reconstruction. This study validated the efficiency of the use of the swab technique to collect DNA for routine parentage assignment, while limiting the adverse effects of destructive tissue sampling for commercial use and/or for the development of breeding programs.