Genetic Characterization of Bay Laurel (Laurus nobilis L.) Populations Using Microsatellite Markers and Flow Cytometry

Abstract

Turkey is one of the few countries that exports the highest quality bay laurel leaf and covers about 90% of the world bay laurel leaf trade. In this study, 95 bay laurel genotypes selected from flora of Hatay province for their superior characteristics were used. Selected genotypes were genetically characterized by 6 SSR markers and the DNA contents were determined by Flow Cytometry. No polyploidy was determined as a result of flow cytometry analysis and 2C DNA values were observed between 5.91 and 6.36 pg. As a result of the SSR analysis, a total of 82 alleles were obtained with a mean of 16.4 of 5 polymorphic loci, while LnD106 loci were observed monomorphic. The highest number of alleles (24 bp) was observed in the LnA2 locus. Generally, a low similarity is determined among the genotypes. The highest genetic similarity was seen in E6 and O6 genotypes with 80%. This situation revealed the importance of genetic diversity in Hatay bay laurel population. The results are important as regard to reveal and protect the genetic diversity of bay laurel existence in Hatay.

Keywords

• Genetic diversity,L. nobilis,DNA content,SSR

Defne (Laurus nobilis L.) Populasyonlarının Mikrosatellit Markörler ve Flow Sitometri ile Genetik Karakterizasyonu

Öz

Türkiye, yüksek kaliteli defne yaprağı ihraç eden birkaç ülkeden biridir ve dünya defne yaprağı ticaretinin yaklaşık% 90'ını Türkiye yapmaktadır. Bu çalışmada üstün özellikleri nedeniyle Hatay florasından seçilen 95 adet defne genotipi kullanılmıştır. Seçilen genotipler, genetik olarak 6 SSR markörü ile karakterize edilmiş ve DNA içerikleri Flow Cytometry ile belirlenmiştir. Flow sitometri analizi sonucunda poliploidi saptanmamış ve 2C DNA değerleri 5.91 ile 6.36 pg arasında gözlenmiştir. SSR analizi sonucunda, 5 polimorfik lokusta ortalama 16.4 ile toplam 82 allel elde edilirken, LnD106 lokusu monomorfik olarak gözlenmiştir. En yüksek allel sayısı (24 bp) LnA2 loküsünde gözlenmiştir. Genel olarak, genotipler arasında benzerlik düşük olmuştur. En yüksek genetik benzerlik oranı % 80 ile E6 ve O6 genotiplerinde görülmüştür Bu durum Hatay defne populasyonundaki genetik çeşitliliğin önemini ortaya çıkarmıştır. Sonuçlar, Hatay'da defne varlığının genetik çeşitliliğini ortaya koymak ve korumak açısından önemlidir. 

Anahtar Kelimeler

• Genetik çeşitlilik,L. nobilis,DNA içeriği

Kaynakça

1. Aboel-Atta AMI, 2009. On the taxonomy of Laurus L. (Lauraceae), evidence from isozymes, RAPD and ISSR. Academic Journal of Plant Sciences 2 (2): 82-91.
2. Acheampong AK, Akromah R and Ofori FA, 2008. Genetic characterization of Ghanaian avocados using microsatellite markers. J. Am Soc Hortic Sci 133(6): 801-809.
3. Alcaraz ML and Hormaza JI, 2007. Molecular characterization and genetic diversity in an avocado collection of cultivars and local Spanish genotypes using SSRs. Hereditas 144 (6): 244-253.
4. Anonymous, 2014. Ormancılık İstatistikleri 2012. Ankara, Türkiye: Türkiye İstatsitik Kurumu Matbaası.
5. Anonymous, 2016. Defne Eylem Planı 2016-2020. Ankara, Türkiye: Orman ve Su İşleri Bakanlığı. Orman Genel Müdürlüğü.
6. Arroyo-Garcia R, Martinez-Zapater JM, Fernandez Prieto JA and Alvarez-Arbesu R, 2001. AFLP evoluation of genetic similarity among laurel populations (Laurus L.). Euphytica 122: 155-164.
7. Arroyo JM, Rigueiro C, Rodriguez R, Hampe A, Valido A, Rodriguez-Sanchez F and Jordano P, 2010. Isolation and characterization of 20 microsatellite loci for laurel species (Laurus, Lauraceae). Am J Bot 97: 26–30.
8. Ayanoğlu F, Kaya DA, Mert A and Köse E, 2013. Determination of quality aspects and selection of native grown laurel (Laurus nobilis L.) in Hatay province of Turkey. The First Mediterranean Symposium on Medicinal and Aromatic Plants (MESMAP) April 17-20, 2013. Gazimagosa, Turkish Republic of Northern Cyprus. p. 59.

9. Bandelj D, Jakse J and Javornik B, 2004. Assessment of genetic variability of olive varieties by microsatellite and AFLP markers. Euphytica 136: 93-102.
10. Baytop T, 1999. Türkiye’de Bitkiler ile Tedavi, Geçmişte ve Bugün 2. Baskı. İstanbul, Türkiye: Nobel Tıp Kitabevleri.
11. Bennett MD and Leitch IJ, 2011. Nuclear DNA amounts in angiosperms: targets, trends and tomorrow. Ann Bot 107: 467–590.
12. Borrone JW, Schnell RJ, Violi H and Ploetz C, 2007. Seventy microsatellite from Persea americana Miller (avokado) express sequence tags. Mol Ecol Notes 7: 439-444.
13. Bowcock AM, Ruiz–Linares A, Tomfohrde J, Minch E, Kidd JR and Cavalli–Sforza LL, 1994. High resolution of human evolutionary trees with polymorphic microsatellites. Nature 368:455-457.
14. Burge GK, Morgan ER and Seelye JE, 2002. Opportunities for synthetic plant chimeral breeding: past and future. Plant Cell, Tissue and Organ Culture 70: 13-21.
15. Ceylan A and Özay N, 1990. Defne yaprakların (Folia lauri)'da ontogenetiksel kalite araştırması. E.Ü.Z.F. Dergisi 27: 71-77.
16. Christenhusz MJM and Byng JW, 2016. The number of known plants species in the world and its annual increase. Phytotaxa 261 (3): 201–217.
17. Davis PH, 1982. Flora of Turkey, Vol. 7. Edinburgh: Edinburgh University Press.
18. Davis J, Henderson D, Kobayashi M, Clegg MT and Cleeg MT, 1998. Genealogical relationships among cultivated avocado as revealed through RFLP analyses. J Hered 89: 319–323.
19. Doyle JJ and Doyle JL, 1987. A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin 19: 11-15.
20. Fiedler J, Bufler G and Bangerth F, 1998. Genetic relationships of avocado (Persea americana Mill.) using RAPD markers. Euphytica 101:249–255.
21. Galbraith DW, 2004. Cytometry and plant sciences: A personal retrospective. Journal of the International Society for Advancement of Cytometry. 58A (1):37–44.
22. Heywood VH, 1978. Flowering Plants of the World. Oxford University Press.
23. Hocquigny S, Pelsy F, Dumas V, Kindt S, Heloir MC and Merdinoglu D, 2004. Diversification within grapevine cultivars goes through chimeric states. Genome 47(3): 579-589.
24. Kayacık H, 1977. Orman ve park agaçlarının özel sistematigi: 2. Angiospermae (Kapalı Tohumlular). İstanbul, Türkiye: İstanbul Univ. Orman Fak. Yayınları.
25. Kekelidze NA, Dzhanikashvili MI and Kutateladze VV, 1987. Dynamics of accumulation and composition formation of essential oil in Laurus nobilis L. leaves during ontogenesis. Fiziol Biokhi Kult 19 (6): 607- 614.
26. Kılıç A, Hafızoğlu H, Kollmannsberger H and Nitz S, 2004. Volatile constituents and key odorants in leaves, buds, flowers and fruits of Laurus nobilis L. J Agr Food Chem 52: 1601-1606.
27. Kumar S, Singh J and Sharma A, 2003. Bay Leaves. In: Peter, KV, Editör. Handbook of Herbs and Spices. Vol. I. Abington Woodhead Publishing Limited, pp. 52-61.
28. Kurt R, Karayılmazlar S, İmren E and Çabuk Y, 2016. Türkiye ormancılık sektöründe odun dışı orman ürünleri: ihracat analizi. Journal of Bartın Faculty of Forestry, 18 (2): 158-167.
29. Lefort F, Lally M, Thompson D and Douglas GC, 1998. Morfological traits microsatellite fingerprinting and genetic relatedness of a stand of elite oaks (Q. robur L.) at Tuallynally, Ireland. Silvae Genet 47: 257-262.
30. Marzouki H, Nasri N, Jouaud B, Bonnet C, Khaldi A, Bouzid S and Fady B, 2009. Population genetic structure of Laurus nobilis L. inferred from transferred nuclear microsatellites. Silvae Genet 58 (5–6): 270-276.
31. Mhameed S, Sharon D, Hillel J, Lahav E, Kaufman D and Lavi U, 1996. Level of heterozygosity and mode of inheritance of variable number of tandem repeat loci in avocado. J Am Soc Hortic Sci 121: 778 - 782.
32. Mhameed S, Sharon D, Kaufman D, Lahav E, Hillel J, Degani C and Lavi U, 1997. Genetic relationships within avocado (Persea americana Mill.) cultivars and between Persea species. Theor Appl Genet 94: 279–286.
33. Minch E, Ruiz-Linares A, Goldstein DB, Feldman M and Cavalli-Sforza LL, 1995. Microsat (version 1.4d): A computer program for calculating various statistics on microsatellite allele data. Stanford. CA, USA:University of Stanford.
34. Paetkau D, Calvert W, Stirling I and Strobeck C, 1995. Microsatellite analysis of population structure in Canadian polar bears. Mol Ecol 4: 347-354.
35. Powell W, Morgante M, Andre C, Hanafey M, Vogel J, Tingey S and Rafalski A, 1996. The comparison of RFLP, RAPD AFLP and SSR (microsatellite) markers for germplasm analysis. Molecular Breeding 2(3): 225-238.
36. Rodilla JM, Tinoco MT, Morais JC, Gimenez C, Cabrera R, Benito DM, Castillo L and Gonzalez-Coloma A, 2008. Laurus novocanariensis essential oil: Seasonal variation and valorization. Biochem Syst Ecol 36: 167-176.
37. Rodriguez-Sanchez F, Guzman B, Valido A, Vargas P and Arroyo J, 2009. Late neogene history of the laurel tree (Laurus L., Lauraceae) based on phylogeographical analyses of Mediterranean and Macaronesian populations. J Biogeogr 36: 1270–1281.
38. Rohlf FJ, 1988. NTSYS-PC: Numerical Taxonomy and Multivariate Analysis System. Version 1.50. New York, USA: Exeter publishing Ltd. & Applied Biostatistics. Inc.
39. Ross IA, 2001. Medicinal Plants of the World Chemical Constituents, Traditional and Modern Medicinal Uses. Vol., 2. New York, USA: Springer Science+Business Media.
40. Sayyah M, Saroukhani G, Peirovi A and Kamalinejad M, 2003. Analgesic and antiinflammatory activity of the leaf essential oil of Laurus nobilis Linn. Phytotherapy Research 17: 733-736.
41. Sefc KM, Lefort F, Grando MS, Scott KD, Steinkellner H and Thomas MR, 2001. Microsatellite markers for grapevine: a state of the art. In Molecular Biology & Biotechnology of the Grapevine (pp. 433-463). Springer Netherlands.
42. Selli F, Bakır M, İnan G, Aygün H, Boz Y, Yaşasın AS, Özer C, Akman B, Söylemezoğlu G, Kazan K and Ergül A, 2007. Simple sequence repeat-based assessment of genetic diversity in 'Dimrit' and 'Gemre' grapevine accessions from Turkey. Vitis 46 (4): 182–187.
43. Simic M, Kundakovic T and Kovacevic N, 2003. Preliminary assay on the antioxidative activity of Laurus nobilis extracts. Fitoterapia. 74 (6): 613-616.
44. Sneath PHA and Sokal RR, 1973. Numerical taxanomy. San Francisco, CA: Freeman.
45. Suda J, Kyncl T and Freiova R, 2003. Nuclear DNA amounts in Macaronesian angiosperms. Ann Bot-London 92: 153-164.
46. Şafak I and Okan T, 2004. Kekik, defne ve çam fıstığının üretimi ve pazarlaması. Doğu Akdeniz Ormancılık Araştırma Müdürlüğü, DOA Dergisi (Journal of DOA) 10: 101-129.
47. Tuna M, Vogel KP, Arumuganathan K and Gill KS, 2001. DNA content and ploidy determination of bromegrass germplasm accessions by flow cytometry. Crop Sci 41: 1629-1634.
48. Wagner HW and Sefc KM, 1999. Identity 1.0. Centre for Applied Genetics, University of Agricultural Science, Vienna.
49. Verdian-Rizi M, 2008. Phenological variation of Laurus nobilis L. essential oil from Iran. EJEAFChe 7: 3321–3325.
50. Zonneveld BJM, Leitch IJ and Bennett MD, 2005. First nuclear DNA amounts in more than 300 angiosperms. Ann Bot-London 96: 229–244.