Reliability of non-invasive tissue sampling methods for DNA extraction in rabbits (Oryctolagus cuniculus)

Manel Ben Larbi, A. Tircazes, K. Feve, F. Tudela, G. Bolet

Abstract

Deoxyribonucleic acid (DNA) can be extracted from different tissue sources. The most common is blood, but in some situations it can be easier to take a biopsy. In some cases when it is difficult to capture animals, especially in wild populations, faeces and hairs can be considered as a source of DNA. This paper presents a pilot study conducted to compare the applicability of invasive and non-invasive sampling methods for extracting DNA for use in genetic studies of rabbits (Oryctolagus cuniculus). The study included 24 rabbits from the INRA 1001 strain.  Blood, hair, ear biopsies and faeces were collected and used as DNA sources. Our aim was to verify the quantity of DNA obtained from different tissues using two or three types of extraction. DNA was obtained for all tissue types and all extraction methods.

DNA extraction was shown to be optimal with the LGC (Laboratory of Cellular Genetics) blood extraction method. With regard to non-invasive methods, DNA extraction for hair using the LGC protocol and QIAamp® DNA mini kit gave very low quantities of DNA that could not be used for PCR reactions. The Chelex extraction protocol gave good results for PCR but could not be quantified. DNA extracted from faeces is a viable source of DNA for determining individual genotypes. The use of such non-invasive samples as a source of genetic material is a recent and very promising technique, especially for the study of endangered species, but these techniques are still too unreliable and costly to altogether replace invasive techniques when the latter are possible.


Keywords

rabbit; DNA extraction; tissue sampling; faeces; hair

Full Text:

PDF

References

Archie E.A., Maldonado J.E., Hollister-Smith J.A., 2008. Fine-scale population genetic structure in a fission-fusion society. Mol Ecol., 17:2666-2679. https://doi.org/10.1111/j.1365-294X.2008.03797.x

Baldwin H.J., Hoggard S.J., Snoyman S.T., Stow A.J., and Brown C., 2010. Non-invasive genetic sampling of faecal materiel and hair from the grey-headed flying-fox (Pteropus poliocephalus). Australian Mammalogy, 32(1): 56-61. https://doi.org/10.1071/AM09015

Ben Larbi M., Haddad B., Allalout S., 2008. Characterization of traditional rabbit breeding system used in the south of Tunisia, 9th world rabbit congress- June 10-13, 2008-Verona-Italy.

Fernando PT, Vidya NC, Pajapakse C, Dangolla A, and Melnick DJ, 2003. Reliable noninvasive genotyping: fantasy or reality. J. Hered., 94: 115-123. https://doi.org/10.1093/jhered/esg022

Fontanesi L., Tazzoli M., Russo V,2007. Non-invasive and simple methodsfor sampling rabbit DNA for PCR analysis of melanocortin 1 receptor (MCR) gene mutations: a technical note. World Rabbit Sci., 15:121-126. https://doi.org/10.4995/wrs.2007.598

Frantzen M.A.J., Silk J.B., Ferguson J.W.H., Wayne R.K. and Kohn M.H. 1998. Empirical evaluation of preservation methods for faecal DNA. Molecular Ecology, 7: 1423-1428. https://doi.org/10.1046/j.1365-294x.1998.00449.x

Gallan M., Baltzinger C., Hewison A.J.M., Cosson.J.F, 2005. Distinguishing red and roe deer using DNA extracted from hair samples and the polymerase chain reaction method. Deer management and research 33(1): 204-211. https://doi.org/10.2193/0091-7648(2005)33[204:DRARDU]2.0.CO;2

Green M.L., Herzing D.L., and Balldwin J.D., 2007. Noninvasive methodology for the sampling and extraction of DNA from free-ranging Atlantic spotted dolphins (Stenella frontalis). Molecular Ecology Notes, 7: 1287-1292. https://doi.org/10.1111/j.1471-8286.2007.01858.x

Kohn M.K., Wayne R.K.,1997. Facts from faeces revisited. Trends in Ecology and Evolution 12:223-227. https://doi.org/10.1016/S0169-5347(97)01050-1

Lindhal T., 1993. Instability and decay of the primary structure of DNA. Nature, 362 : 709-715. https://doi.org/10.1038/362709a0

Mainnguy J., Bernatchez L., 2007. L'analyse de l'ADN sans manipulation des animaux : un outil incontournable pour la gestion et la conservation des espèces rares et élusives. Le naturaliste canadien, 31: 51-59.

Miller S.A., Dykes D.D., Polesky H.F., 1988. A sampling salting out procedure for extracting DNA from human nucleated cells. Nucleic acids research 16: 1215. https://doi.org/10.1093/nar/16.3.1215

Morin P.A., Chambers K.E., Boesch C.H. and Vigilant L., 2001. Quantitative polymerase chain reaction analysis of DNA from non-invasive samples for accurate microsatellite genotyping of wild chimpanzees (Pan troglodytes verus). Molecular Ecology, 10 : 1835-1844. https://doi.org/10.1046/j.0962-1083.2001.01308.x PMid:11472550

Mougel F., Mounolou J-C.and Monnerot M., 1997. Nine polymorphic microsatellite loci in the rabbit, Oryctolagus cuniculus. Animal genetics 28: 58-71. https://doi.org/10.1111/j.1365-2052.1997.00047.x

Roon, D.A., Waits L.P. and Kendall K.C., 2003. A quantitative evaluation of two methods for preserving hair samples. Molecular Ecology Notes, 3 : 163-166. https://doi.org/10.1046/j.1471-8286.2003.00358.x

Taberlet, P., Griffin S., Goossens B., Questiau S., Manceau V., . Escaravage N, . Waits L.P, and Bouvet J., 1996. Reliable genotyping of samples with very low DNA quantities using PCR. Nucleic Acids Research, 24: 3189-3194. https://doi.org/10.1093/nar/24.16.3189

Wehausen J.D., Ramey II R.R. and Epps C.W., 2004. Experiments in DNA extraction and PCR amplification from Bighorn sheep faeces: The importance of DNA extraction methods. Journal of Heredity, 95(6):503-509. https://doi.org/10.1093/jhered/esh068

Abstract Views

3152
Metrics Loading ...

Metrics powered by PLOS ALM




 

 Universitat Politècnica de València

 

Official journal of the World Rabbit Science Association (WRSA)

 

e-ISSN: 1989-8886     ISSN: 1257-5011   https://doi.org/10.4995/wrs