Development and characterization of microsatellite markers for the endangered Laotian rock-rat ( Laonastes aenigmamus )
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TECHNICAL NOTE
Development and characterization of microsatellite markers for the endangered Laotian rock-rat (Laonastes aenigmamus) using 454-sequencing technology Jorge Luis Pino • Marina Sofia Ascunce David Reed • Jean-Pierre Hugot
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Received: 31 May 2012 / Accepted: 23 June 2012 / Published online: 8 July 2012 Ó Springer Science+Business Media B.V. 2012
Abstract The Laotian rock rat (Laonastes aenigmamus) is the single surviving member of the family Diatomyidae, which has a distribution restricted to the karstic region of Lao-PDR. Here we describe the development of 12 polymorphic microsatellites markers for the endangered Laotian rock rat using 454-sequencing. We successfully tested 12 markers in 30 individuals from 2 populations. Eleven of the 12 loci were polymorphic and the number of alleles detected per locus ranged from 2 to 11. Three of these loci deviated significantly from Hardy–Weinberg equilibrium, which coincides with the detection of possible null alleles. These microsatellite markers are expected to contribute in future research and conservation of L. aenigmamus. Keywords Laotian rock rat Laonastes aenigmanus Khanyou Microsatellite 454 sequencing Conservation genetics
Since the discovery of the Laotian rock rat or kha-nyou (Laonastes aenigmamus) in 2005 (Jenkins et al. 2005), several studies have described the taxonomy of the species and genus while others using molecular and morphological data have examined its phylogenetic placement into the family Diatomyidae (Jenkins et al. 2005; Dawson et al. 2006; Huchon et al. 2007). Regardless, this enigmatic J. L. Pino (&) M. S. Ascunce D. Reed Florida Museum of Natural History, Dickinson Hall, Museum Rd & Newell Dr., Gainesville, FL 32611, USA e-mail: [email protected] J.-P. Hugot Muse´um National d’Histoire Naturelle, De´partement Syste´matique et Evolution, UMR CNRS 7205, 47 rue Cuvier, CP 51, Paris, France
rodent is the single surviving member of its family and investigations at the population level have not been made in fine scale. Laonastes has a narrow distribution in a karstic region located primarily in the Lao People’s Democratic Republic, in Southeast Asia (Jenkins et al. 2005). Classified as endangered by the UICN, its populations are considered to be declining due to habitat loss and hunting (Aplin and Lunde 2008). Recent molecular data suggest that this species has strong geographical structure (Rivie`re-dobigny et al. 2011) highlighting the necessity for the development of microsatellite markers to aid in assessing species status definitions and fine-scale genetic differentiation. Here we describe 12 polymorphic microsatellite markers designed for L. aenigmamus using 454-sequencing. Whole genomic DNA was isolated from Laonastes liver tissue using standard DNeasy Blood and Tissue kit protocol (QIAGEN). Genomic sample from one individual was submitted, to the University of Florida ICBR, for sequencing on one-eighth of a plate using 454GS-FLX technology (Roche Applied Science). We obtained 92,433 fragment reads from the 454-sequ
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