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Australian Journal of Zoology Australian Journal of Zoology Society
Evolutionary, molecular and comparative zoology
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RESEARCH ARTICLE
Contents Vol 57(4)

Development of microsatellite markers for the short-beaked echidna using three different approaches

C. Vanpé A B , E. Buschiazzo B , J. Abdelkrim A B , G. Morrow C , S. C. Nicol C and N. J. Gemmell A B D
+ Author Affiliations
- Author Affiliations

A Centre for Reproduction and Genomics, Department of Anatomy and Structural Biology, University of Otago, PO Box 56, Dunedin 9054, New Zealand.

B Molecular Ecology Laboratory, School of Biological Science, University of Canterbury, Private Bag 4800, Christchurch, New Zealand.

C School of Zoology, University of Tasmania, Private Bag 5, Hobart, Tasmania 7001, Australia.

D Corresponding author. Email: neil.gemmell@otago.ac.nz

Australian Journal of Zoology 57(4) 219-224 https://doi.org/10.1071/ZO09033
Submitted: 2 April 2009  Accepted: 3 June 2009   Published: 26 October 2009

Abstract

We used three different methods, size-selected genomic library, cross-species amplification of a mammal-wide set of conserved microsatellites and genomic sequencing, to develop a panel of 43 microsatellite loci for the short-beaked echidna (Tachyglossus aculeatus). These loci were screened against 13 individuals from three different regions (Tasmania, Kangaroo Island, Perth region), spanning the breadth of the range of the short-beaked echidna. Nine of the 43 tested loci amplified reliably, generated clear peaks on the electropherogram and were polymorphic, with the number of alleles per locus ranging from two to eight (mean = 3.78) in the individuals tested. Polymorphic information content ranged from 0.16 to 0.78, and expected heterozygosity ranged from 0.19 to 0.84. One of the nine microsatellites showed a heterozygote deficit, suggesting a high probability of null alleles. The genomic sequencing approach using data derived from the Roche FLX platform is likely to provide the most promising method to develop echidna microsatellites. The microsatellite markers developed here will be useful tools to study population genetic structure, gene flow, kinship and parentage in Tachyglossus sp. and potentially also in endangered Zaglossus species.

Additional keywords: 454 sequencing, cross-species amplification, echidna, genomic library, microsatellite, monotremes, Tachyglossus aculeatus.


Acknowledgements

We acknowledge Brita Hansen for her initial work on the genomic library and Michelle French for her help in genotyping. We thank Peggy Rismiller and Arthur Ferguson of the Perth zoo for providing echidna tissue samples. Tom Pringle provided information on echidna 454 reads. Jo-Ann Stanton converted the sff files of 454 sequencing reads into Fasta format. CV was supported by a Lavoisier postdoctoral research fellowship from the French Ministry of Foreign and European Affairs. We gratefully acknowledge support from the National Geographic Committee for Research and Exploration and the University of Canterbury for funding and logistical support.


References

Abdelkrim, J. , Robertson, B. C. , Stanton, J. A. L. , and Gemmell, N. J. (2009). Fast, cost-effective development of species-specific microsatellite markers by genomic sequencing. BioTechniques 46(3), 185–192.
| Crossref | GoogleScholarGoogle Scholar | CAS | PubMed | Augee M. , Gooden B. , and Musser A. (2006). ‘Echidna: Extraordinary Egg-laying Mammal.’ (CSIRO Publishing: Melbourne.)

Benjamini, Y. , and Hochberg, Y. (1995). Controlling the false discovery rate: a practical and powerful approach to multiple testing. Journal of the Royal Statistical Society. Series B. Methodological 57, 289–300.
Buschiazzo E. (2008). Conservation and evolution of microsatellites in vertebrate genomes. Ph.D. Thesis, University of Canterbury, Christchurch, New Zealand.

Buschiazzo, E. , and Gemmell, N. J. (2009). Evolutionary and phylogenetic significance of platypus microsatellites conserved in vertebrate genomes. Australian Journal of Zoology 57, 175–184.
| Crossref | GoogleScholarGoogle Scholar | Griffiths M. (1968). ‘Echidnas.’ (Pergamon Press: Oxford.)

Griffiths M. (1978). ‘The Biology of the Monotremes.’ (Academic Press: New York.)

Grützner, F. , Deakin, J. , Rens, W. , El-Mogharbel, N. , and Marshall Graves, J. A. (2003). The monotreme genome: a patchwork of reptile, mammal and unique features? Comparative Biochemistry and Physiology. Part A, Molecular & Integrative Physiology 136(4), 867–881.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Moritz C. , Wilmer J. W. , Pope L. , Sherwin W. B. , Taylor A. C. , and Limpus C. J. (1996). Applications of genetics to the conservation of Australian fauna: four case studies from Queensland. In ‘Molecular Genetic Approaches in Conservation’. (Eds T. B. Smith and R. K. Wayne.) pp. 442–456. (Oxford University Press: Oxford.)

Morrow, G. , Andersen, N. A. , and Nicol, S. C. (2009). Reproductive strategies of the short-beaked echidna – a review with new data from a long-term study on the Tasmanian subspecies (Tachyglossus aculeatus setosus). Australian Journal of Zoology 57, 275–282.
| Crossref | GoogleScholarGoogle Scholar | Sambrook J. , Fritsch E. F. , and Maniatis T. (1989). ‘Molecular Cloning: A Laboratory Manual.’ (Cold Spring Harbor: New York.)

Schuelke, M. (2000). An economic method for the fluorescent labeling of PCR fragments. Nature Biotechnology 18, 233–234.
| Crossref | GoogleScholarGoogle Scholar | CAS | PubMed |

Storey, J. D. (2002). A direct approach to false discovery rates. Journal of the Royal Statistical Society. Series B. Methodological 64, 479–498.
| Crossref | GoogleScholarGoogle Scholar |

Temple-Smith, P. D. , and Grant, T. (2001). Uncertain breeding: a short history of reproduction in monotremes. Reproduction, Fertility and Development 13(8), 487–497.
| Crossref | GoogleScholarGoogle Scholar | CAS |

Tóth, G. , Gáspári, Z. , and Jurka, J. (2000). Microsatellites in different eukaryotic genomes: survey and analysis. Genome Research 10, 967–981.
| Crossref | GoogleScholarGoogle Scholar | PubMed |

Walsh, P. S. , Metzger, D. A. , and Higuchi, R. (1991). Chelex® 100 as a medium for simple extraction of DNA for PCR-based typing from forensic material. BioTechniques 10(4), 506–513.
|  CAS | PubMed |

Warren, W. C. , Hillier, L. W. , Graves, J. A. M. , Birney, E. , and Ponting, C. P. , et al. (2008). Genome analysis of the platypus reveals unique signatures of evolution. Nature 455(7210), 256.
| Crossref | GoogleScholarGoogle Scholar | CAS |

Watson, J. M. , Riggs, A. D. , and Graves, J. A. M. (1992). Gene mapping studies confirm the homology between the platypus X and echidna X1 chromosomes and identify a conserved ancestral monotreme X. Chromosoma 101, 596–601.
| Crossref | GoogleScholarGoogle Scholar | CAS | PubMed |

Wrigley, J. M. , and Graves, J. A. M. (1988). Karyotypic conservation in the mammalian order Monotremata (subclass Prototheria). Chromosoma 96, 231–247.
| Crossref | GoogleScholarGoogle Scholar | CAS | PubMed |

Zane, L. , Bargelloni, L. , and Patarnello, T. (2002). Strategies for microsatellite isolation: a review. Molecular Ecology 11, 1–16.
| Crossref | GoogleScholarGoogle Scholar | CAS | PubMed |

Zenger, K. R. , Eldridge, M. D. B. , Pope, L. C. , and Cooper, D. W. (2003). Characterisation and cross-species utility of microsatellite markers within kangaroos, wallabies and rat kangaroos (Macropoidea: Marsupialia). Australian Journal of Zoology 51, 587–596.
| Crossref | GoogleScholarGoogle Scholar | CAS |