Twenty-five novel microsatellite markers for English sole, Parophrys vetulus
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MICROSATELLITE LETTERS
Twenty-five novel microsatellite markers for English sole, Parophrys vetulus Gary A. Winans • Jon D. Baker • Stacey L. Lance
Received: 21 November 2013 / Accepted: 25 November 2013 Ó Springer Science+Business Media Dordrecht (outside the USA) 2013
Abstract Although English sole Parophrys vetulus has been used extensively as a bioindicator of anthropogenic pollution in the US Pacific Northwest, little is known about its genetic population structure. We isolated and characterized a total of 25 microsatellite loci from 23 individuals from Whidbey Island, Washington. The number of alleles per locus ranged from 5 to 19, observed heterozygosity ranged 0.091–0.957, and the probability of identity values ranged from 0.009 to 0.157. These new loci will be used to study genetic variability within and among populations in the Salish Sea (Puget Sound and Strait of Georgia) to help monitor effects of near shore urbanization, and establish and monitor marine protected areas. Keywords Parophrys vetulus Microsatellite Illumina Pal_finder SSR Population markers
The English sole (Pleuronectidae) is a commercially important flatfish distributed from the Bering Sea to Baja, Mexico. It is one of several flatfish species which, along with the spotted ratfish (Hydrolagus colliei), make up the principal fish biomass in the Puget Sound Basin. Although English sole has been studied extensively in the Puget Sound to evaluate physiological, metabolic, and behavioral
G. A. Winans (&) NOAA Northwest Fisheries Science Center, Seattle, WA 98112, USA e-mail: [email protected] J. D. Baker Douglas Salmometrics, Mukilteo, WA 98275, USA S. L. Lance Savannah River Ecology Laboratory, Aiken, SC 29802, USA
responses to anthropogenic pollutants (Johnson et al. 2008), little is known about its genetic population structure. Understanding stock structure of a key benthic marine animal will help monitor near shore urbanization and establish effective marine protected areas. Genomic DNA was extracted and purified from caudal fin clips using the Qiagen DNeasy 96 Blood and Tissue kit, spin-column protocol (Qiagen, Valencia, California, USA). Following extraction, the sample was quantified with a Qubit 2.0 Fluorometer (Life Technologies, Grand Island, New York, USA1) using the Qubit dsDNA BR Assay Kit (Life Technologies, Grand Island, New York, USA). Two samples were standardized to 60 ng/ll and 23 samples were standardized to 20 ng/ll. An Illumina paired-end shotgun library was prepared by shearing 1 lg of DNA using a Covaris S220 and following the standard protocol of the Illumina TruSeq DNA Library Kit and using a multiplex identifier adaptor index. Illumina sequencing was conducted on the HiSeq with 100 bp paired-end reads. Five million of the resulting reads were analyzed with the program PAL_FINDER_v0.02.03 (Castoe et al. 2012) to extract those reads that contained di-, tri-, tetra-, penta-, and hexanucleotide microsatellites. Once positive reads were identified in PAL_FINDER_v0.02.03 they were batched to a local in
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