Abstract

In this research, determination of phylogenetic tree of sheep in Şanlıurfa province using molecular techniques was the main goal. White Karaman (AK) and Awassi sheep (IV) breed raised in Şanlıurfa province were used as the animal materials. Fleece samples were collected for genomic DNA isolation in sheep, and genomic DNAs were isolated in all the samples. In DNA samples, necessary forward and reverse primers were designed to amplify mitochondrial Cytochrome b (Cyt b) gene region. Mitochondrial Cyt b gene region were amplified by applying polymerase chain reaction (PCR) technique, and gene sequence information of PCR products were obtained. The rate of G+C, number of polymorphic site (S), number of haplotypes (h), haplotype diversity (Hd), nucleotide diversity (π) and total number of region for all populations were calculated. N-J phylogenetic tree formed in this research using sheep haplotype Cyt b sequences and reference sequences ( for A, B, C, D and E lineage), 6 haplotypes (IV01, AK13, AK09, IV06, AK16, IV05) out of 16 haplotypes were in B lineage, 1 haplotype (IV12) were in A lineage, 1 haplotype (AK04) were in E lineage, 5 haplotypes (AK12, AK02, IV16, AK01, IV08, and IV03) were in C lineage, and AK06 and IV13 were in different group. In conclusion, in White Karaman and Awassi sheep, raised in Şanlıurfa province; gene sequences of Cytochrome b gene were determined.

Şanlıurfa Yöresi Akkaraman ve İvesi Koyunlarında Mitokondriyal Sitokrom b (Cyt b) Gen Dizisine Göre Filogenetik Analizler

Abstract

Bu çalışmada, Şanlıurfa yöresindeki Akkaraman (AK) ve İvesi (IV) koyunlarının filogenetik yapıları moleküler tekniklerle belirlenmeye çalışılmıştır. Araştırmanın hayvan materyalini, Şanlıurfa ve yöresinde yetiştirilen Akkaraman (AK) ve İvesi (IV) koyunları oluşturmuştur. Koyunlardan genomik DNA izolasyonu için kıl örnekleri toplanmış ve tüm örneklerden genomik DNA izole edilmiştir. Koyun DNA örneklerinde mitokondriyal Sitokrom b (Cyt b) gen bölgelerini çoğaltmak için gerekli ileri ve geri primerler tasarlanmıştır. Koyun Cyt b gen bölgeleri polimeraz zincir reaksiyonu tekniği (PZR) ile çoğaltılmıştır. PZR ürünlerinin gen dizi bilgileri elde edilmiştir. Populasyonlar için toplam bölge sayısı, G+C oranı, polimorfik bölge sayısı (S), haplotip sayısı (h), haplotip farklılığı (Hd) ve nükleotid farklılığı (π) değerleri hesaplanmıştır. Koyun haplotiplere ait Cyt b gen dizileri ile referans diziler (A, B, C, D, ve E soyları) birlikte oluşturulan N-J filogenetik ağaçta, 16 haplotipten, 6’sı B soyunda (IV01, AK13, AK09, IV06, AK16, IV05), 1’i A soyunda (IV12), 1’i E soyunda (AK04) 5’i C soyunda (AK12, AK02, IV16, AK01, IV08, IV03) yer almış, AK06 ve IV13 farklı kümelenmiştir. Sonuç olarak, Şanlıurfa yöresi Akkaraman ve İvesi koyunları Cyt b gen bölgesi gen dizileri belirlenmiştir.

Keywords

• Koyun
• Filogenetik
• mtDNA
• Moleküler çalışmlar

References

1. Bradley, D.G., Machugh, D.E., Cunningham. P., Loftus, R.T. 1996. Mitochondrial diversity and the origins of african and european cattle. Proceedings of the National Academy of Sciences of the United States of America, 93 (10): 5131-5135.
2. Guo, J., Du, L.X., Ma, Y.H., Guan, W.J., Li, H.B., Zhao, Q.J., Li, X., Rao, S.Q. 2005. A novel maternal lineage revealed in sheep (Ovis aries). Animal Genetics, 36 (4): 331-336.
3. Hiendleder, S., Lewalski, H., Wassmuth, R., Ke, A. 1998a. The complete mitochondrial DNA sequence of the domestic sheep (Ovis aries) and comparison with the other major ovine haplotype. Journal of Molecular Evolution, 47 (4): 441-448.
4. Hiendleder, S., Mainz, K., Plante, Y., Lewalski, H. 1998b. Analysis of mitochondrial dna indicates that domestic sheep are derived from two different ancestral maternal sources: no evidence for contributions from Urial and Argali sheep. Journal of Heredety, 89 (2): 113-120.
5. Hiendleder, S., Phua, Sh., Hecht, W. 1999. A diagnostic assay discriminating between two major Ovis aries mitochondrial DNA haplogroups. Animal Genetics, 30 (3): 211-213.
6. Hiendleder, S., Kaupe, B., Wassmut, R., and Janke, A. 2002. Molecular analysis of wild and domestic sheep qestions current nomenclature and provides evidence for domestication from two different subspecies. Proceedings of The Royal Society of London, 269: 893-904.
7. Joshi, M.B., Rout, P.K., Mandal, A.K., Tyler-Smith, C., Singh, L., Thangaraj, K. 2004. Phylogeography and origin of Indian domestic goats. Molecular Biology Evolution, 21 (3): 454-462.
8. Kımura, M. 1980. A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution, 16: 111-120.

9. Librado, P., Rozas, J. 2009. DnaSP v5: A software for comprehensive analysis of DNA polymorphism data. Bioinformatics, 25: 1451-1452.
10. Loftus, R. T., Machugh, D. E., Bradley, D. G., Sharp, P. M., Cunningham, P. 1994. Evidence for two independent domestications of cattle. Proceedings of the National Academy of Sciences of the United States of America, 91: 2757–2761.
11. Luikart, G., Gielly, L., Excoffier, L., Vigne, Jd., Bouvet, J., Taberlet, P. 2001. Multiple maternal origins and weak phylogeographic structure in domestic goats. Proceedings of the National Academy of Sciences of the United States of America, 98 (10): 5927-5932.
12. Machugh, D.E., Bradley, D.G. 2001. Livestock genetic origins: goats buck the trend. Proceedings of the National Academy of Sciences of the United States of America, 98 (10): 5382-5384.
13. Mannen, H., Nagata, Y., Tsujı, S. 2001. Mitochondrial DNA reveal that domestic goat (Capra hircus) are genetically affected by two subspecies of benzoar (Capra aegagurus). Biochemical Genetics, 39 (5-6): 145-54.
14. Meadows, J.R., Cemal, I., Karaca, O., Gootwine, E., Kijas, J.W. 2007. Five ovine mitochondrial lineages identified from sheep breeds of the near East. Genetics, 175 (3): 1371-1379. Nei, M., Kumar, S. 2000. Molecular evolution and phylogenetics, Oxford University Press, Oxford.
15. Pardeshi, V.C., Kadoo, N.Y., Sainani M.N., Meadows, J.R., Kijas, J.W., Gupta, V.S. 2007. Mitochondrial haplotypes reveal a strong genetic structure for three Indian sheep breeds. Animal Genetics, 38 (5): 460-466.
16. Pedrosa, S., Uzun, M., Arranz, Jj., Gutierrez-Gil, B., San, Primitivo, F., Bayon, Y. 2005. Evidence of three maternal lineages in near eastern sheep supporting multiple domestication events. Proceedings of The Royal Society B, 272 (1577): 2211-2217.
17. Pedrosa, S., Arranz, J.J., Brito, N., Molia, A., San Primitivo, F., Bayón, Y. 2007. Mitochondrial diversity and the origin of Iberian sheep. Genetics Selection Evolution, 39 (1): 91-103. Pereira, F., Davis, S.J., Pereira, L., Mcevoy, B., Bradley, D.G., Amorim, A. 2006. Genetic signatures of a Mediterranean influence in Iberian Peninsula sheep husbandry. Molecular Biology and Evolution, 23 (7): 1420-1426.
18. Ryder, M.L. 1984. Sheep. In: Evolution of domesticated animals, Longman, London, 63-85.
19. Saitou, N., Nei, M. 1987. The neighbor-joining method: a new method for reconstruction phylogenetic trees. Molecular Biology and Evolution, 4: 406-425.
20. Sambrook, J., Fritsch, E. F., Maniatis, T. 1989. Molecular cloning:A laboratory manual (2nd ed.), 3 vol., Cold-Spring Harbor, New York.
21. Sultana, S., Mannen, H., Tsuji, S. 2003. Mitochondrial DNA diversity of Pakistani goats. Animal Genetics, 34 (6): 417-421.
22. Tamura, K., Dudley, J., Nei, M., Kumar, S. 2007. MEGA4: Molecular evolutionary genetics analysis (MEGA) software version 4.0. Molecular Biology and Evolution, 24:1596-1599. Tapio, M., Marzanov, N., Ozerov, M.., Cinkulov, M., Gonzarenko, G., Kiselyova, T., Murawskıi M., Vinalass, H., Kantanen , J. 2006. Sheep mitochondrial DNA variation in European, Caucasian, and Central Asian areas. Molecular Biology and Evolution, 23 (9): 1776-1783.
23. Thompson, J.D., Higgins, D.G., Gibson, T.J., 1994. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice. Nucleic Acids Research, 22: 4673-4680.
24. Troy, C.S., Machugh, D.E., Bailey, J.F., Magee, D.A., Loftus, R.T., Cunningham, P., Chamberlain, A.T., Sykes, B.C., Bradley, D.G. 2001. Genetic evidence for Near-Eastern origins of European cattle. Nature, 410: 1088–1091.
25. Zeder, Ma., Hesse, B. 2000 . The initial domestication of goats (Capra Hircus) in the Zagros mountains 10.000 years ago. Science, 287 (5461): 2254-2257.
26. Zeuner, F.E. 1963. A history of domesticated animals. Harper & Row: New York.