Development and characterization of sixteen microsatellite markers for the federally endangered species: Leptodea leptod
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TECHNICAL NOTE
Development and characterization of sixteen microsatellite markers for the federally endangered species: Leptodea leptodon (Bivalvia: Unionidae) using paired-end Illumina shotgun sequencing Jason O’Bryhim • Jer Pin Chong • Stacey L. Lance Kenneth L. Jones • Kevin J. Roe
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Received: 19 March 2012 / Accepted: 2 April 2012 / Published online: 18 April 2012 Ó Springer Science+Business Media B.V. 2012
Abstract We isolated and characterized a total of 16 microsatellite loci in Leptodea leptodon. Loci were screened in 24 individuals from across the species current range in Missouri. The number of alleles per locus ranged from 3 to 17, observed heterozygosity ranged from 0.208 to 1.000, and the probability of identity values ranged from 0.014 to 0.67. These new loci will be used for conducting investigations into the genetic structure and diversity of extant populations of this federally endangered species. Keywords Leptodea Microsatellite PAL_FINDER PCR primers SSR
The scaleshell Leptodea leptodon (Rafinesque, 1820) is a federally endangered species (USFWS 2001). Whereas this species once occurred in 56 rivers in 13 states within the Mississippi River Basin, it is now restricted to 13 streams in Missouri, Arkansas and Oklahoma, with the largest known populations occurring in the Meramec, Bourbeuse and
J. O’Bryhim S. L. Lance Savannah River Ecology Laboratory, University of Georgia, Aiken, SC 29802, USA J. O’Bryhim Department of Environmental Science and Policy, George Mason University, Fairfax, VA 22030, USA J. P. Chong K. J. Roe (&) Department of Natural Resource Ecology and Management, Iowa State University, Ames, IA 50010-3221, USA e-mail: [email protected] K. L. Jones Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA
Gasconade rivers in Missouri (USFWS 2010). Degradation of water quality, sedimentation, channel destabilization, and habitat destruction are all considered to be contributing factors in the decline of the scaleshell throughout its range. Very little is known about the life history, reproductive biology and the extant population size of L. leptodon (USFWS 2010), and the development of species specific genetic markers are consistent with the goals outlined in the USFWS recovery plan, including improving our understanding of the biology and ecology of the scaleshell mussel. Total DNA was extracted from one individual of Leptodea leptodon, following the protocol of Puregene Buccal Cell DNA Kit (Gentra Systems, Inc. Minneapolis, MN) for use in isolation of microsatellite loci. 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. This library was pooled with those from other species and 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 (Ca
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