NOFSal-MP10: A single hypervariable STR 12-plex for accurate verification of triploidy in Atlantic salmon (Salmo salar L.)

Abstract

An increasing interest in production of sterile fish in aquaculture requires rapid, accurate and efficient testing for certification of triploidy prior to sale. Further, validation of triploidy can be beneficial, and even required, for accrediting methods of triploid production. A PCR-genotyping approach using a single megaplex of 12 hypervariable STR markers shows accurate and highly repeatable results enabling verification of ploidy in a single test. The NOFSal-MP10 panel contains 12 STR markers mapped to 9 chromosomes with an average of 21.4 alleles per marker for a combined total of 257 alleles based on genotyped samples to-date. The hypervariable nature of these 12 STRs leads to a large probability for three uniquely sized alleles to be observed at each marker, thus providing a rapid confirmation of ploidy based on the count of allele fragments per marker. Further, as a PCR amplification step is involved, this method is robust to DNA quality and quantity, making it suitable for very early determination of ploidy, as early as the eyed-egg stage. Repeat genotyping of positive control diploid Atlantic salmon over two different capillary electrophoretic instruments in different laboratories and with multiple laboratory personnel proves the panels robustness to scoring errors with an overall allelic error rate of 0.3% and a false-positive triploid assignment rate of zero. Genotyping of DNA from 1238 eggs and larvae from 18 independent triploid production batches over three years confirmed triploidy in 98% of samples based on a semi-strict criterion of three unique alleles at one or more loci, and 95% based on a strict criterion of three unique alleles at two or more loci. HighlightsO_LIThis paper describes a rapid quantifiable test for validation of triploidy in Atlantic salmon C_LIO_LIThe method is highly accurate and repeatable and robust to DNA quality, allowing testing in embryonic stages C_LIO_LIRapid early testing of triploidy will enable certified sales of triploid eggs for production C_LI