Development of microsatellite loci for the Honduran white-bat ( Ectophylla alba ) by using Illumina paired-end sequences
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Development of microsatellite loci for the Honduran white-bat (Ectophylla alba) by using Illumina paired-end sequences Alberto Morales-Leyva • Rodrigo A. Medellı´n • Stacey L. Lance • Bernal Rodrı´guez-Herrera • Melina Del Real-Monroy • Jorge Ortega
Received: 28 August 2013 / Accepted: 23 September 2013 / Published online: 17 October 2013 Ó Springer Science+Business Media Dordrecht 2013
Abstract Ectophylla alba is a bat restricted to Costa Rica, Honduras, Nicaragua and Panama´. A technique based on Illumina paired-end sequencing of a library highly enriched for microsatellite repeats was used to develop loci. Thirteen polymorphic (tri, tetra- and pentanucleotide) microsatellites were developed and tested. All markers were genotyped on 42 different individuals from 5 distinct locations. We observed low to medium–high genetic variation across most loci. Levels of expected heterozygosity across all markers was medium to low (mean HE = 0.659, mean HO = 0.672). Keywords Ectophylla alba Illumina Microsatellites Pal_finder
A. Morales-Leyva M. Del Real-Monroy J. Ortega (&) Laboratorio de Bioconservacio´n y Manejo, Posgrado en Ciencias Quimicobiolo´gicas, Departamento de Zoologı´a, Escuela Nacional de Ciencias Biolo´gicas, Instituto Polite´cnico Nacional, Prolongacio´n de Carpio y Plan de Ayala s/n, Col. Sto. Tomas, 11340 Mexico City, Mexico e-mail: [email protected] R. A. Medellı´n Laboratorio de Ecologı´a y Conservacio´n de Vertebrados, Departamento de Ecologı´a de la Biodiversidad, Instituto de Ecologı´a, Universidad Nacional Auto´noma de Me´xico, Circuito Exterior junto a Jardı´n Bota´nico, 04510 Mexico City, Mexico S. L. Lance Savannah River Ecology Laboratory, University of Georgia, Aiken, SC 29803, USA B. Rodrı´guez-Herrera Seccio´n de Zoologı´a, Escuela de Biologı´a, Universidad de Costa Rica, San Pedro Montes de Oca, San Jose´, Costa Rica
Ectophylla alba is a small, distinctively white bat, with a forearm length among 27–31 mm (Rodrı´guez-Herrera et al. 2006). The white tent-making bat has a small geographic range restricted to the Caribbean lowlands of Honduras, Costa Rica, Nicaragua and Panama´ (Rodrı´guez-Herrera et al. 2006). It is considered a lower to near threatened species by the IUCN. Wing punches were collected and kept in alcohol (96 %), from 42 wild individuals of E. alba from five different localities of Costa Rica. DNA was extracted following instructions of the Qiagen protocol (Blood and Tissue Kit, Cat No. 69504). 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. 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 installation of the program Primer3 (version 2.0.0) for primer design. We recovered 3180
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