, Volume 135, Issue 3, pp 457–470 | Cite as

Assaying polymorphism at DNA level for genetic diversity diagnostics of the safflower (Carthamus tinctorius L.) world germplasm resources

  • Deepmala Sehgal
  • Vijay Rani Rajpal
  • Soom Nath Raina
  • Tsuneo Sasanuma
  • Tetsuo Sasakuma


Carthamus tinctorius (2n = 2x = 24), commonly known as safflower, is widely cultivated in agricultural production systems of Asia, Europe, Australia, and the Americas as a source of high quality vegetable and industrial oil. Twenty-two RAPD primers, 18 SSR primers, and 10 AFLP primer combinations were used to assess: (1) the genetic diversity of 85 accessions (originating from 24 countries) representing global germplasm variability of safflower and (2) the interrelationships among safflower ‘centers of similarity’ or ‘regional gene pools’ proposed earlier. The RAPD and SSR primers and AFLP primer combinations revealed 57.6, 68.0, and 71.2% polymorphism, respectively, among 111, 72, and 330 genetic loci amplified from the accessions. The sum of effective number of alleles (66.44), resolving power (59.16), and marker index (51.3) explicitly revealed the relative superiority of AFLP as a marker system in uncovering variation in safflower. Overall, AFLP markers could recognize ‘centers of similarity’ or ‘regional gene pools’. Analysis of molecular variance and Shannon’s information index provided corroborating evidences for the present and previous studies that concluded fragmentation of safflower gene pool into many gene pools. Divergent directional selection is likely to have played an important role in shaping the diversity. From the practical applications standpoint, the diversity of Iran–Afghanistan gene pool is very high, equivalent to the total diversity of the species. The Far East gene pool is the least diverse. The present comprehensive input, first of its own kind in safflower, will assist marker based improvement programmes in the crop.


AFLP Carthamus tinctorius L. Gene pools Genetic diversity ISSR RAPD 



Thanks are due to the United States Department of Agriculture (USDA) for supplying seed samples. This work was supported, in part, by Department of Biotechnology and Council of Scientific and Industrial Research, Ministry of Science and Technology, Government of India, and by the special coordination funds of Science and Technology Agency of the Japan Government.


  1. Amel SH, Khaled C, Messaoud M, Mohamed M, Mokhtar T (2005) Comparative analysis of genetic diversity in two Tunisian collections of fig cultivars based on random amplified polymorphic DNA and inter simple sequence repeats fingerprints. Genet Resour Crop Evol 52:563–573. doi: 10.1007/s10722-003-6096-3 CrossRefGoogle Scholar
  2. Ashri A (1971a) Evaluation of world collection of safflower C. tinctorius L. I. Reaction to several diseases and association with morphological characters in Israel. Crop Sci 11:253–257Google Scholar
  3. Ashri A (1971b) Evaluation of world collection of C. tinctorius L. II. Resistance to safflower fly A. helianthi R. Euphytica 20:410–415. doi: 10.1007/BF00035666 CrossRefGoogle Scholar
  4. Ashri A (1973) Divergence and evolution in the safflower genus Carthamus L. Final research report, PL 480, USDA, The Hebrew University of Jerusalem, Faculty of Agriculture, Rehovot, IsraelGoogle Scholar
  5. Ashri A (1975) Evaluation of the germplasm collection of safflower Carthamus tinctorius L. V. Distribution and regional divergence for morphological characters. Euphytica 24:651–659. doi: 10.1007/BF00132903 CrossRefGoogle Scholar
  6. Ashri A, Knowles PF, Urie AL, Zimmer DE, Cahaner A, Marani A (1975) Evaluation of the germplasm collection of safflower Carthamus tinctorius III Oil content and iodine value and their associations with other characters. Econ Bot 31:38–46Google Scholar
  7. Ashri A, Zimmer DE, Urie AL, Cahaner A, Marani A (1974) Evaluation of world collection of safflower Carthamus tinctorius L. IV Yield and yield components and their relationships. Crop Sci 14:799–802Google Scholar
  8. Aslam M, Hazara GR (1993) Evaluation of world collection of safflower (Carthamus tinctorius L.) for yield and other agronomic characters In: Dajue L, Yuanzhou H (eds) Third international safflower conference, Beijing, China, 9–13 June 1993, p 238Google Scholar
  9. Bornet B, Goraguer F, Joly G, Branchard M (2002) Genetic diversity in European and Argentinian cultivated potatoes (Solanum tuberosum subsp tuberosum) detected by inter-simple sequence repeats (ISSRs). Genome 45:481–484. doi: 10.1139/g02-002 PubMedCrossRefGoogle Scholar
  10. Bussel JD (1999) The distribution of random amplified polymorphic DNA (RAPD) diversity amongst populations of Isotoma petraea (Lobeliaceae). Mol Ecol 8:775–789. doi: 10.1046/j.1365-294X.1999.00627.x CrossRefGoogle Scholar
  11. Carapetian J, Estilai A (1997) Genetics of isozyme coding genes in safflower. In: Corleto A, Mundel HH (eds) Proceedings of the 4th International safflower conference: Safflower: a multipurpose species with unexploited potential and world adaptability, Adriatica, Editrice, Bari, Italy, 2–7 June 1997, pp 235-237Google Scholar
  12. Charlesworth B, Nordborg M, Charlesworth D (1997) The effects of local selection, balanced polymorphism, and background selection on equilibrium patterns of genetic diversity in sub-divided populations. Genet Res 70:155–174. doi: 10.1017/S0016672397002954 PubMedCrossRefGoogle Scholar
  13. Dellaporta SL (1994) Plant DNA miniprep and microprep: versions 2.1–2.3. In: Freeling M, Walbolt V (eds) The maize handbook. Springer New York Inc., New York, pp 522–525Google Scholar
  14. Efron Y, Peleg M, Ashri A (1973) Alcohol dehydrogenase allozymes in the safflower genus Carthamus L. Biochem Genet 9:299–308. doi: 10.1007/BF00485742 PubMedCrossRefGoogle Scholar
  15. Excoffier L, Smouse P, Quattro J (1992) Analysis of molecular variance inferred from metric distances among DNA haplotypes: application to human mitochondrial DNA restriction data. Genetics 131:479–491PubMedGoogle Scholar
  16. Fernandez-Martinez J, Rio M, Haro A (1993) Survey of safflower (Carthamus tinctorius L.) germplasm for variants in fatty acid composition and other seed characters. Euphytica 69:115–122CrossRefGoogle Scholar
  17. Fischer M, Husi R, Prati D, Peintinger M, van Kleunen M, Schmid B (2000) RAPD variation among and within small and large populations of the rare clonal plant Ranunculus reptans (Ranunculaceae). Am J Bot 87:1128–1137. doi: 10.2307/2656649 PubMedCrossRefGoogle Scholar
  18. Futehally S (1982) Inheritance of very high levels of linoleic acid in the seed oil of safflower (Carthamus tinctorius L.). MS thesis, University of California, DavisGoogle Scholar
  19. Genet T, Viljoen CD, Labuschagne MT (2005) Genetic analysis of Ethiopian mustard genotypes using amplified fragment length polymorphism (AFLP) markers. Afr J Biotechnol 4:891–897Google Scholar
  20. Ghebru B, Schmidt RJ, Bennetzen JL (2002) Genetic diversity of Eriterian sorghum landraces assessed with simple sequence repeat (SSR) markers. Theor Appl Genet 105:229–236. doi: 10.1007/s00122-002-0929-x PubMedCrossRefGoogle Scholar
  21. Han Y, Li D (1992) Evaluation of safflower (Carthamus tinctorius L.) germplasm–analysis in fatty acid composition of seeds of domestic and exotic safflower varieties. Bot Res 6:28–35Google Scholar
  22. Hanelt P (1961) Zur Kenntnis von Carthamus tinctorius L. Kulturpflanze 9:114–145. doi: 10.1007/BF02095747 (in German)CrossRefGoogle Scholar
  23. He G, Prakash CS (1997) Identification of polymorphic DNA markers in cultivated peanuts (Arachis hypogaea L.). Euphytica 97:143–149. doi: 10.1023/A:1002949813052 CrossRefGoogle Scholar
  24. Hongtrakul V, Huestis GM, Knapp SJ (1997) Amplified fragment length polymorphisms as a tool for DNA fingerprinting sunflower germplasm: genetic diversity among oilseed inbred lines. Theor Appl Genet 95:400–407. doi: 10.1007/s001220050576 CrossRefGoogle Scholar
  25. Howard A, Howard GL, Khan AR (1910) The economic significance of natural cross-fertilization in India. Mem Dept Agric India. Bot Ser 3:281–330Google Scholar
  26. Kadam BS, Patrankar VK (1942) Natural cross-pollination in safflower. Indian J Genet Plant Breed 2:69–70Google Scholar
  27. Keim P, Beavis W, Schupp J, Freestone R (1992) Evaluation of soybean RFLP marker diversity in adapted germplasm. Theor Appl Genet 85:205–212. doi: 10.1007/BF00222861 CrossRefGoogle Scholar
  28. Keim P, Maschinski J, Travis SE (1996) An analysis of genetic variation in Astragalus cremnophylax var cremnophylax, a critically endangered plant, using AFLP markers. Mol Ecol 5:735–745. doi: 10.1111/j.1365-294X.1996.tb00370.x PubMedCrossRefGoogle Scholar
  29. Knowles PF (1958) Safflower. Adv Agron 10:289–323. doi: 10.1016/S0065-2113(08)60068-1 CrossRefGoogle Scholar
  30. Knowles PF (1968) Associations of high levels of oleic acid in the seed oil of safflower (Carthamus tinctorius) with other plant and seed characteristics. Econ Bot 22:195–200Google Scholar
  31. Knowles PF (1969a) Modification of quantity and quality of safflower oil through plat breeding. J Am Oil Chem Soc 46:130–132. doi: 10.1007/BF02635715 CrossRefGoogle Scholar
  32. Knowles PF (1969b) Centers of plant diversity and conservation of crop germplasm: safflower. Econ Bot 23:324–329Google Scholar
  33. Knowles PF (1972) The plant geneticist’s contribution toward changing lipid and amino acid composition of safflower. J Am Oil Chem Soc 49:27–29. doi: 10.1007/BF02545133 CrossRefGoogle Scholar
  34. Ladd SL, Knowles PF (1970) Inheritance of stearic acid in the seed oil of safflower (Carthamus tinctorius L.). Crop Sci 10:525–527Google Scholar
  35. Le Clerc V, Briard M, Revollon P (2002) Influence of number and map distribution of AFLP markers on similarity estimates in carrot. Theor Appl Genet 106:157–162PubMedGoogle Scholar
  36. Lima MLA, Garcia AAF, Oliviera KM, Matsuoko S, Arizono H, de Sonza CL Jr et al (2002) Analysis of genetic similarity detected by AFLP and coefficient of parentage among genotypes of sugarcane (Saccharum spp). Theor Appl Genet 104:30–38. doi: 10.1007/s001220200003 PubMedCrossRefGoogle Scholar
  37. Lübberstedt T, Melchinger AE, Dussle C, Vuylsteke M, Kuiper M (2000) Relationships among early European maize inbreds: IV. Genetic diversity revealed with AFLP markers and comparison with RFLP, RAPD and pedigree data. Crop Sci 40:783–791Google Scholar
  38. Mantel N (1967) The detection of disease clustering and a generalized regression approach. Cancer Res 27:209–220PubMedGoogle Scholar
  39. Marsan PA, Castiglioni P, Fusari K, Kuiper M, Motto M (1998) Genetic diversity and its relationship to hybrid performance in maize as revealed by RFLP and AFLP markers. Theor Appl Genet 96:219–227. doi: 10.1007/s001220050730 CrossRefGoogle Scholar
  40. Milbourne D, Meyer R, Bradshaw JE, Baird E, Bonar N, Provan J et al (1997) Comparison of PCR-based marker systems for the analysis of genetic relationships in cultivated potato. Mol Breed 3:127–136. doi: 10.1023/A:1009633005390 CrossRefGoogle Scholar
  41. Negi MS, Sabharwal V, Bhat SR, Lakshmikumaran M (2004) Utility of AFLP markers for the assessment of genetic diversity within Brassica nigra germplasm. Plant Breed 123:13–16. doi: 10.1046/j.0179-9541.2003.00926.x CrossRefGoogle Scholar
  42. Pejic I, Ajmone-Marsan P, Morgante M, Kozumplick V, Castiglioni P, Taramino G et al (1998) Comparative analysis of genetic among maize inbred lines detected by RFLPs, RAPDs, SSRs and AFLPs. Theor Appl Genet 97:1248–1255. doi: 10.1007/s001220051017 CrossRefGoogle Scholar
  43. Perera L, Rusell JR, Provan J, McNicol JW, Powell W (1998) Evaluating genetic relationships between indigenous coconut (Cococ nucifera L.) accessions from Sri Lanka by means of AFLP profiling. Theor Appl Genet 96:545–550. doi: 10.1007/s001220050772 CrossRefGoogle Scholar
  44. Portis E, Barchi L, Acquadro A, Macua JI, Lanteri S (2005) Genetic diversity assessment in cultivated cardoon by AFLP (amplified fragment length polymorphism) and microsatellite markers. Plant Breed 124:299–304. doi: 10.1111/j.1439-0523.2005.01098.x CrossRefGoogle Scholar
  45. Powell W, Morgante M, Andre C, Hanafey M, Vogel J, Tingey S et al (1996) The comparison of RFLP, RAPD, AFLP and SSR (microsatellite) markers for germplasm analysis. Mol Breed 2:225–238. doi: 10.1007/BF00564200 CrossRefGoogle Scholar
  46. Prevost A, Wilkinson MJ (1999) A new system of comparing PCR primers applied to ISSR fingerprinting of potato cultivars. Theor Appl Genet 98:107–112. doi: 10.1007/s001220051046 CrossRefGoogle Scholar
  47. Rohlf FJ (1998) NTSYS-pc: numerical taxonomy and multivariate analysis system version 2.02K Applied Biostatistics, New YorkGoogle Scholar
  48. Schut JW, Qi X, Stam P (1997) Association between relationship measures based on AFLP markers, pedigree data and morphological traits in barley. Theor Appl Genet 95:161–1168. doi: 10.1007/s001220050677 CrossRefGoogle Scholar
  49. Sehgal D, Bhat V, Raina SN (2008a) Advent of DNA markers to decipher genome sequence polymorphism. In: Kirti PB (ed) Handbook of new technologies for genetic improvement of grain legumes, CRC Press, New York, pp 477–495Google Scholar
  50. Sehgal D, Bhat V, Raina SN (2008b) Applicability of DNA markers for genome diagnostics of grain legumes. In: Kirti PB (ed) Handbook of new technologies for genetic improvement of grain legumes, CRC Press, New York, pp 497–557Google Scholar
  51. Sehgal D, Raina SN (2005) Genotyping safflower (Carthamus tinctorius L.) cultivars by DNA fingerprints. Euphytica 146:67–76. doi: 10.1007/s10681-005-8496-2 CrossRefGoogle Scholar
  52. Sehgal D, Raina SN (2008c). DNA markers and germplasm resource diagnostics: new perspectives in crop improvement and conservation strategies. In: Arya ID, Arya S (eds) Utilization of biotechnology in plant sciences. Rastogi Press, Meerut, India, pp 39–54Google Scholar
  53. Soleimani VD, Baum BR, Johnson DA (2002) AFLP and pedigree-based genetic diversity estimates in modern cultivars of durum wheat Triticum turgidum L.subsp. durum (Desf.) Husn. Theor Appl Genet 104:350–357. doi: 10.1007/s001220100714 PubMedCrossRefGoogle Scholar
  54. Souframanien J, Gopalakrishna T (2004) A comparative analysis of genetic diversity in blackgram genotypes using RAPD and ISSR markers. Theor Appl Genet 109:1687–1693. doi: 10.1007/s00122-004-1797-3 PubMedCrossRefGoogle Scholar
  55. Staub JE, Danin-Poleg Y, Fazio G, Horejsi T, Reis N, Katzir N (2000) Comparative analysis of cultivated melon groups (Cucumis melo L.) using random amplified polymorphic DNA and simple sequence repeat markers. Euphytica 115:225–241. doi: 10.1023/A:1004054014174 CrossRefGoogle Scholar
  56. Steinger T, Haldimann P, Leiss KA, Müller-Schärer H (2002) Does natural selection promote population divergence? a comparative analysis of population structure using amplified fragment length polymorphism markers and quantitative traits. Mol Ecol 11:2583–2590. doi: 10.1046/j.1365-294x.2002.01653.x PubMedCrossRefGoogle Scholar
  57. Tam SM, Mhiri C, Vogelaar A, Kerkveld M, Pearce SR, Grandbastien A (2005) Comparative analyses of genetic diversities within tomato and pepper collections detected by retrotransposons-based SSAP, AFLP and SSR. Theor Appl Genet 110:819–831. doi: 10.1007/s00122-004-1837-z PubMedCrossRefGoogle Scholar
  58. Thein SI, Wallace RR (1986) The use of synthetic oligonucleotides as specific hybridization probes in the diagnosis of genetic disorders. In: Davis KE (ed) Human genetic diseases: a practical approach. IRL, Oxford, pp 33–50Google Scholar
  59. Tosti N, Negri V (2005) On-going on-farm microevolutionary processes in neighbouring cowpea landraces revealed by molecular markers. Theor Appl Genet 110:1275–1283. doi: 10.1007/s00122-005-1964-1 PubMedCrossRefGoogle Scholar
  60. Ude G, Pillay M, Ogundiwin E, Tenkouano A (2003) Genetic diversity in an African plantain core collection using AFLP and RAPD markers. Theor Appl Genet 107:248–255. doi: 10.1007/s00122-003-1246-8 PubMedCrossRefGoogle Scholar
  61. Uptmoor R, Wenzel W, Friedt W, Donaldson G, Ayisi K, Ordon F (2003) Comparative analysis on the genetic relatedness of Sorghum bicolor accessions from Southern Africa by RAPDs, AFLPs and SSRs. Theor Appl Genet 106:1316–1325PubMedGoogle Scholar
  62. Virk PS, Zhu J, Newbury HJ, Bryan GJ, Jackson MT, Ford-Llyod BV (2000) Effectiveness of different classes of molecular marker for classifying and revealing variation in rice (Oryza sativa) germplasm. Euphytica 112:275–284. doi: 10.1023/A:1003952720758 CrossRefGoogle Scholar
  63. Weiss EA (1971) Castor, sesame and safflower. Leonard Hill Books/University Press, Aberdeen, London, pp 529–774Google Scholar
  64. Weiss EA (1983) Oilseed crops. Longman Group Limited, London, pp 216–281Google Scholar
  65. Yee E, Kidwell KK, Sills GR, Lumpkin TA (1999) Diversity among selected Vigna angularis (Azuki) accessions on the basis of RAPD and AFLP markers. Crop Sci 39:268–275Google Scholar
  66. Zeid M, Schön C, Link W (2003) Genetic diversity in recent elite faba bean lines using AFLP markers. Theor Appl Genet 107:1304–1314. doi: 10.1007/s00122-003-1350-9 PubMedCrossRefGoogle Scholar
  67. Zhao J, Wang X, Deng B, Lou P, Wu J, Sun R et al (2005) Genetic relationships within Brassica rapa as inferred from AFLP fingerprints. Theor Appl Genet 110:1301–1314. doi: 10.1007/s00122-005-1967-y PubMedCrossRefGoogle Scholar
  68. Zhebentyayeva TN, Reighard GL, Gorina VM, Abbott AG (2003) Simple sequence repeat (SSR) analysis for assessment of genetic variability in apricot germplasm. Theor Appl Genet 106:435–444PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Deepmala Sehgal
    • 1
    • 2
  • Vijay Rani Rajpal
    • 3
  • Soom Nath Raina
    • 3
  • Tsuneo Sasanuma
    • 1
  • Tetsuo Sasakuma
    • 1
  1. 1.Kihara Institute for Biological ResearchYokohama City UniversityYokohamaJapan
  2. 2.Institute of Grassland and Environmental Research (IGER)Aberystwyth UniversityAberystwythUK
  3. 3.Laboratory of Cellular and Molecular Cytogenetics, Department of BotanyUniversity of DelhiDelhiIndia

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