Using eggshell membranes as a DNA source for population genetic research

Abstract

In the context of population genetic research, a faster and less invasive method of DNA sampling would allow large-scale assessments of genetic diversity and genetic differentiation with the help of volunteer observers. The aim of this study was to investigate the usefulness of eggshell membranes as a DNA source for population genetic research, by addressing eggshell membrane DNA quality, degeneration and cross-contamination. To this end, a comparison was made with blood-derived DNA samples. We have demonstrated 100% successful DNA extraction from post-hatched Black-tailed Godwit (Limosa limosa) eggshell membranes as well as from blood samples. Using 11 microsatellite loci, DNA amplification success was 99.1% for eggshell membranes and 97.7% for blood samples. Genetic information within eggshell membrane DNA in comparison to blood DNA was not affected (F ST = −0.01735, P = 0.999) by degeneration or possible cross-contamination. Furthermore, neither degeneration nor cross-contamination was apparent in total genotypic comparison of eggshell membrane DNA and blood sample DNA. Our research clearly illustrates that eggshell membranes can be used for population genetic research.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2

References

  1. Bowkett AE, Plowman AB, Stevens JR, Davenport TRB, van Vuuren BJ (2009) Genetic testing of dung identification for antelope surveys in the Udzungwa Mountains, Tanzania. Conserv Genet 10:251–255. doi:10.1007/s10592-008-9564-7

    CAS  Article  Google Scholar 

  2. Bush KL, Vinsky MD, Aldridge CL, Paszkowski CA (2005) A comparison of sample types varying in invasiveness for use in DNA sex determination in an endangered population of greater Sage-Grouse (Centrocercus uropihasianus). Conserv Genet 6:867–870. doi:10.1007/s10592-005-9040-6

    Article  Google Scholar 

  3. Coulon A, Fitzpatrick JW, Bowman R, Stith BM, Makarewich CA, Stenzler LM, Lovette IJ (2008) Congruent population structure inferred from dispersal behaviour and intensive genetic surveys of the threatened Florida scrub-jay (Aphelocoma coerulescens). Mol Ecol 17:1685–1701. doi:10.1111/j.1365-294X.2008.03705.x

    CAS  Article  Google Scholar 

  4. Excoffier LGL, Schneider S (2005) Arlequin ver. 3.0: an integrated software package for population genetics data analysis. Evol Bioinform Online 1:47–50

    CAS  Article  Google Scholar 

  5. Hoglund J, Johansson T, Beintema A, Schekkerman H (2009) Phylogeography of the black-tailed Godwit Limosa limosa: substructuring revealed by mtDNA control region sequences. J Ornithol 150:45–53. doi:10.1007/s10336-008-0316-8

    Article  Google Scholar 

  6. Kalinowski ST (2002) How many alleles per locus should be used to estimate genetic distances? Heredity 88:62–65. doi:10.1038/sj.hdy.6800009

    CAS  Article  Google Scholar 

  7. Kalinowski ST (2005) Do polymorphic loci require large sample sizes to estimate genetic distances? Heredity 94:33–36. doi:10.1038/sj.hdy.6800548

    CAS  Article  Google Scholar 

  8. Larsson JK, Jansman HAH, Segelbacher G, Hoglund J, Koelewijn HP (2008) Genetic impoverishment of the last black grouse (Tetrao tetrix) population in the Netherlands: detectable only with a reference from the past. Mol Ecol 17:1897–1904. doi:10.1111/j.1365-294X.2008.03717.x

    Article  Google Scholar 

  9. Liebezeit JR, Smith PA, Lanctot RB, Schekkerman H, Tulp I, Kendall SJ, Tracy DM, Rodriques RJ, Meltofte H, Robinson JA, Gratto-Trevor C, McCaffery BJ, Morse J, Zack SW (2007) Assessing the development of shorebird eggs using the flotation method: species-specific and generalized regression models. Condor 109:32–47. doi:10.1650/0010-5422(2007)109[32:ATDOSE]2.0.CO;2

    Article  Google Scholar 

  10. Manier MK, Arnold SJ (2005) Population genetic analysis identifies source-sink dynamics for two sympatric garter snake species (Thamnophis elegans and Thamnophis sirtalis). Mol Ecol 14:3965–3976. doi:10.1111/j.1365-294X.2005.02734.x

    Article  Google Scholar 

  11. Milot E, Weimerskirch H, Bernatchez L (2008) The seabird paradox: dispersal, genetic structure and population dynamics in a highly mobile, but philopatric albatross species. Mol Ecol 17:1658–1673. doi:10.1111/j.1365-294X.2008.03700.x

    CAS  Article  Google Scholar 

  12. Ortego J, Calabuig G, Aparicio J, Cordero PJ (2008) Genetic consequences of natal dispersal in the colonial lesser kestrel. Mol Ecol 17:2051–2059. doi:10.1111/j.1365-294X.2008.03719.x

    Article  Google Scholar 

  13. Ottvall R, Hoglund J, Bensch S, Larsson K (2005) Population differentiation in the redshank (Tringa totanus) as revealed by mitochondria DNA and amplified fragment length polymorphism markers. Conserv Genet 6:321–331. doi:10.1007/s10592-005-4973-3

    CAS  Article  Google Scholar 

  14. Pearce JM, Fields RL, Scribner KT (1997) Nest materials as a source of genetic data for avian ecological studies. J Field Ornithol 68:471–481

    Google Scholar 

  15. Qiagen (2003) DNeaasy Tissue Handbook. protocol for isolation of total DNA from animal tussues. QIAGEN. Vlaencia, California, USA, pp.18-20

  16. Richardson DS, Jury FL, Blaakmeer K, Komdeur J, Burke T (2001) Parentage assignment and extra-group paternity in a cooperative breeder: the Seychelles warbler (Acrocephalus sechellensis) Mol Ecol 10:2263–2273

    CAS  Article  Google Scholar 

  17. Schmaltz G, Somers CM, Sharma P, Quinn JS (2006) Non-destructive sampling of maternal DNA from the external shell of bird eggs. Conserv Genet 7:543–549. doi:10.1007/s10592-005-9065-x

    CAS  Article  Google Scholar 

  18. Schroeder J, Lourenco PM, Van der Velde M, Hooijmeijer JCEW, Both C, Piersma T (2008) Sexual dimorphism in plumage and size in black-tailed Godwits Limosa limosa limosa. Ardea 96:25–37

    Article  Google Scholar 

  19. Strausberger BM, Ashley MV (2001) Eggs yield nuclear DNA from egg-laying female cowbirds, their embryos and offspring. Conserv Genet 2:385–390. doi:10.1023/A:1012526315617

    CAS  Article  Google Scholar 

  20. Taberlet P, Fumagalli L (1996) Owl pellets as a source of DNA for genetic studies of small mammals. Mol Ecol 5:301–305. doi:10.1111/j.1365-294X.1996.tb00318.x

    CAS  Article  Google Scholar 

  21. Taberlet P, Waits LP (1998) Non-invasive genetic sampling. Trends Ecol Evol 13:26–27. doi:10.1016/S0169-5347(97)01276-7

    CAS  Article  Google Scholar 

  22. Van den Brink V, Schroeder J, Both C, Lourenco PM, Hooijmeijer JCEW, Piersma T (2008) Space use by Black-tailed Godwits Limosa limosa limosa during settlement at a previous or a new nest location. Bird Study 55:188–193

    Article  Google Scholar 

  23. Van Oosterhout C, Hutchinson WF, Wills DPM, Shipley P (2004) MICRO-CHECKER: software for identifying and correcting genotyping errors in microsatellite data. Mol Ecol Notes 4:535–538. doi:10.1111/j.1471-8286.2004.00684.x

    Article  Google Scholar 

  24. Verkuil YI, Trimbos K, Haddrath O, Baker AJ (2009) Characterization of polymorphic microsatellite DNA markers in the black-tailed godwit (Limosa limosa: Aves). Mol Ecol (in press)

Download references

Acknowledgments

We hereby declare that the experiments comply with the current laws of the country in which they were performed. We would like to thank Theunis Piersma, Jos Hooijmeijer, Petra de Goeij, Pedro Lourenco and the rest of the Groningen University Black-tailed Godwit group for their help in collecting eggshells and blood samples, storing the eggshells and providing laboratory space for DNA extractions. Marco van der Velde from the University of Groningen was of great help with these extractions. Yvonne Verkuil from the Groningen University helped with understanding the Arlequin and Micro-checker software. Klaas Vrieling and Rene Glas of the Leiden Institute of Biology (IBL) were helpful with PCR amplifications and providing laboratory space. We are also grateful to Nigel Harle for brushing up our English.

Open Access

This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Krijn Baptist Trimbos.

Additional information

Communicated by M. Wink.

Rights and permissions

Open Access This is an open access article distributed under the terms of the Creative Commons Attribution Noncommercial License (https://creativecommons.org/licenses/by-nc/2.0), which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

Reprints and Permissions

About this article

Cite this article

Trimbos, K.B., Broekman, J., Kentie, R. et al. Using eggshell membranes as a DNA source for population genetic research. J Ornithol 150, 915–920 (2009). https://doi.org/10.1007/s10336-009-0422-2

Download citation

Keywords

  • Limosa limosa
  • Population genetics
  • Microsatellite
  • Eggshell membrane
  • DNA quality