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Primer Notes
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Characterization of 37 breed-specific single nucleotide polymorphisms in sheep.
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L. Pariset1, I. Cappuccio1, P. Ajmone-Marsan2, M. Bruford3, S. Dunner4, O. Cortes4, G.
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Erhardt5, S. Joost7, G. Juma3, I. Nijman6, J.A.. Lenstra6, T. Perez3, C. Peter5, A. Valentini1
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and Econogene Consortium8
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Dipartimento di Produzioni Animali, Università della Tuscia, Viterbo, Italy
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Istituto di Zootecnica, Università Cattolica del Sacro Cuore, Piacenza, Italy
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School of Biosciences, University of Wales, Cardiff, United Kingdom
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Departimento de Producción Animal, Universidad Complutense, Madrid, Spain
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Department of Animal Breeding and Genetics, Justus-Liebig-Universität, Giessen, Germany
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Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands
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Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
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http://lasig.epfl.ch/projets/econogene
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Abstract
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We characterized 37 single nucleotide polymorphisms (SNPs) in sheep identified using a targeted-
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gene approach within the Econogene project. Inter-breed polymorphisms were identified on
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individuals belonging to 8 different sheep breeds selected throughout Europe. Genotypes of sheep
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from the 8 breeds were determined for all the SNPs characterized and basic statistics were
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calculated for each SNP. So far, there are not sheep SNPs available in NCBI dbSNP database and
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about 100 are reported in literature. Therefore they will be a useful complement to currently
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available sheep markers.
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Keywords
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Sheep, single nucleotide polymorphisms, allele frequency.
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Correspondence: L. Pariset, fax : +39 761 357434, e-mail: pariset@unitus.it
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Single nucleotide polymorphisms (SNPs) are frequent (Brouillette et al. 2000; Shubitowski et al.
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2001), suitable for ecological and conservation studies (Morin et al., 2004; Seddon et al., 2005;
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Vignal et al. 2002) and valuable for estimating parameters such as population history and inference
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of relationships (Kuhner et al., 2000; Glaubitz et al., 2003). Therefore they represent an ideal
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genetic marker for population and evolutionary studies (Sunnucks 2000). SNPs are not employed
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frequently in studies of non-model organisms, primarily due to difficulties in finding SNPs in
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species where little DNA sequence data are available (Aitken et al 2004). Their use could lead to a
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rapid, large scale and cost effective genotyping (Schlotterer, 2004; Syvanen et al., 2001; Vitalis et
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al., 2001; Vignal et al., 2002).
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SNPs were discovered by across-breed comparison of 16 unrelated individuals belonging to 8
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European sheep breeds representing the most of the variation across a wide geographic area:
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Akkaraman (Turkey), Bergamasca (Italy), Karagouniko (Greece), Rhönsheep (Germany), Rubia del
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Molar (Spain), Turcana (Romania), Welsh Mountain (Great Britain), Zelazna. (Poland). DNA was
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isolated after collection of whole blood using standard techniques. Thirty-seven SNPs have been
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identified and genotyped in about 30 individuals of each of the same 8 breeds, sampling no more
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than 3 individuals per farm.
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Primers designed using sheep sequences (where available) or the consensus sequences of the closest
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species in Genebank were used both for PCR amplification and sequencing of the corresponding
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genomic fragment. Sequences were BLASTed to check for homology greater than 90%. Table 1
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reports loci in which SNPs were identified, characterization details and the common population
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statistics (Weir, 1996, Botstein et al., 1980) for each locus and over all populations. Data were
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computed using the programs Powermarker (Liu and Muse, 2001) and Genepop 3.3 (Raymond and
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Rousset, 1995). Table 2 shows the gene diversity (Nei, 1987), the Fis (Weir and Cockerham, 1984)
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and the frequencies of the rare alleles for each locus and each breed, calculated using the program
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Fstat (Goudet, 2000).
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Of the 37 identified SNPs, 31 were transitions, 5 were transversions and 1 was a deletion. 32 SNPs
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show an overall frequency of the rare allele higher than 5%, resulting appropriate for standard
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population genetic analysis. Of the 5 SNPs showing frequencies lower than 5%, some have higher
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frequency in specific breeds. In particular, the frequency of CALPA1 (0.031 overall breeds) is
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0.203 in Karagouniko and 0.132 in Rhönsheep, the frequency of CALPA2 (0.030 overall breeds) is
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0.266 in Turcana and 0.108 in Welsh Mountain, the frequency of LEPTIN1 (0.012 overall breeds)
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is 0.245 in Welsh Mountain and LEPTIN2 (0.024 overall breeds) reaches a frequency of 0.165 in
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Turcana and of 0.245 in Welsh Mountain. Those SNPs can be employed to reconstruct breed’s
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history or are likely to be under selection in those breeds.
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Acknowledgements:
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This work has been partially supported by the EU Econogene contract QLK5-CT-2001-02461. The
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content of the publication does not represent necessarily the views of the Commission or its
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services.
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