TECHNICAL NOTE Fourteen novel microsatellite markers for the gopher frog, Lithobates capito (Amphibia: Ranidae) Schyler O. Nunziata • Stephen C. Richter • Robert D. Denton • John M. Yeiser • Danielle E. Wells • Kenneth L. Jones • Cris Hagen • Stacey L. Lance Received: 10 August 2011 / Accepted: 28 August 2011 Ó Springer Science+Business Media B.V. 2011 Abstract We isolated and characterized a total of 14 microsatellite loci from gopher frogs, Lithobates capito. This species is of conservation concern because most populations have gone locally extinct across the geographic distribution. Loci were screened for 21 individuals from a single population in Florida. The number of alleles per locus ranged from 7 to 17, observed heterozygosity ranged from 0.667 to 0.947, and the probability of identity values ranged from 0.011 to 0.077. These new loci provide tools for examining the genetic diversity and population structure of L. capito populations and addressing factors associated with their decline. Keywords Lithobates Á Rana Á Ranidae Á Gopher frog Á Microsatellite Á PCR primers Á SSR Á STR Gopher frogs, Lithobates capito, are habitat specialists that inhabit periodically burned long-leaf pine (Pinus palustris) ecosystems in the non-reproductive season and migrate to depressional wetlands for reproduction (Jensen and Richter 2005). As a result of habitat loss, L. capito is now con- sidered endangered, threatened, or of special concern in all states they inhabit (Jensen and Richter 2005), and populations in portions of the geographic distribution may warrant federal protection. Currently, there are eight microsatellite loci developed for L. sevosus, dusky gopher frogs, that cross amplify in L. capito (Richter and Broughton 2005), but to increase power of analyses we developed additional loci for this species. Total DNA was extracted from one individual of L. capito, using the DNeasy tissue kit protocol (Qiagen, Valencia, CA) for the construction of the microsatellite library. DNA was then serially enriched twice for micro- satellites using three probe mixes following Glenn and Schable (2005) with changes described in Lance et al. (2010). There were two primary changes to the Glenn and Schable (2005) protocol. First, a different linker was used (SimpleX-5 Forward 5 0 -AAAA CGAGCAGCGGAACT and SimpleX-5 Reverse 5 0 -pAGTTCCGCTGCTCG). Sec- ond, the enriched libraries were sequenced on a 454 using titanium chemistry following standard Roche 454 library protocols (454 Life Sciences, a Roche company, Branford CT). All methods for sequencing, microsatellite identifica- tion, primer design, and primer screening are as described Disclaimer This report was prepared as an account of work sponsored by an agency of the United States Government. Neither the United States Government nor any agency thereof, nor any of their employees, makes any warranty, express or implied, or assumes any legal liability or responsibility for the accuracy, completeness, or usefulness of any information, apparatus, product, or process disclosed, or represents that its use would not infringe privately owned rights. Reference herein to any specific commercial product, process, or service by trade name, trademark, manufacturer, or otherwise does not necessarily constitute or imply its endorsement, recommendation, or favoring by the United States Government or any agency thereof. The views and opinions of authors expressed herein do not necessarily state or reflect those of the United States Government or any agency thereof. S. O. Nunziata Á S. C. Richter (&) Á R. D. Denton Á J. M. Yeiser Á D. E. Wells Department of Biological Sciences, Moore 235, Eastern Kentucky University, 521 Lancaster Avenue, Richmond, KY 40475, USA e-mail: stephen.richter@eku.edu S. O. Nunziata Á C. Hagen Á S. L. Lance Savannah River Ecology Laboratory, University of Georgia, Aiken, SC 29803, USA K. L. Jones University of Colorado School of Medicine, Aurora, CO 80045, USA 123 Conservation Genet Resour DOI 10.1007/s12686-011-9508-2