Horticulture, Environment, and Biotechnology

, Volume 56, Issue 2, pp 216–224 | Cite as

Development of genomic SSR markers and genetic diversity analysis in cultivated radish (Raphanus sativus L.)

  • Kyung-Mi Bae
  • Sung-Chur Sim
  • Jee-Hwa Hong
  • Keun-Jin Choi
  • Do-Hoon Kim
  • Yong-Sham KwonEmail author
Research Report


Radish (Raphanus sativus L.) is a major vegetable cultivated worldwide. As a member of the family Brassicaceae, this species has diverse morphological characteristics in root. Radish cultivars have been classified based on morphological traits, including root shape and color. Despite its economic importance in Asia, genomic research in radish is less well developed relative to Brassica rapa, a close relative of radish. In this study, we developed genomic simple sequence repeat (SSR) markers using a SSR-enriched library and investigated genetic diversity in a collection of 144 radish cultivars. A total of 237 primer pairs for SSRs were designed and 184 (77.6%) primer pairs produced PCR amplicons. Of these, we selected 27 SSR markers (14.7%) based on polymorphism in a subset of 11 cultivars and then used to assess genetic relationships in the germplasm panel. For these markers, the number of alleles per marker ranged from 2 to 18 with an average of 7.77 alleles and the polymorphism information content (PIC) values ranged from 0.491 to 0.906. The estimates of pairwise F st revealed significant genetic differentiation between the five market classes of 135 radish cultivars (74 big, 15 small, 29 young, 12 Altari, and 5 processing). Clustering analysis using NTSYS-pc and STRUCTURE software also found that the 74 big radishes were divided into 5–7 clusters. In addition, all 27 SSR markers were able to differentiate 64 big radish cultivars based on the UPGMA dendrogram and each of the 23 markers independently identified 1 to 17 big radish cultivars. These results suggest that cultivated radishes have different patterns of genetic variation and breeding practices should be a driving force for the genetic differentiation between and within market classes. The SSR markers developed in this study will be a useful resource for genetic study and protection of plant breeder’s intellectual property right through cultivar identification.

Additional key words

molecular marker SSR-enriched library polymorphism clustering analysis genetic differentiation 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Supplementary material

13580_2015_89_MOESM1_ESM.pdf (224 kb)
Supplementary material, approximately 244 KB.

Literature Cited

  1. Anderson, J.A., G.A. Churchill, J.E. Autrique, S.D. Tanksley, and M.E. Sorrells. 1993. Optimizing parental selection for genetic-linkage maps. Genome 36:181–186.CrossRefPubMedGoogle Scholar
  2. Blair, M.W., M.C. Giraldo, H.F. Buendia, E. Tovar, M.C. Duque, and S.E. Beebe. 2006. Microsatellite marker diversity in common bean (Phaseolus vulgaris L.). Theor. Appl. Genet. 113:100–109.CrossRefPubMedGoogle Scholar
  3. Clapham, A.R., T.G. Tutin, and E.F. Warburg. 1962. Flora of the British Isles. Cambridge University Press, Cambridge, UK.Google Scholar
  4. Cloutier, S., Z. Niu, R. Datla, and S. Duguid. 2009. Development and analysis of EST-SSRs for flax (Linum usitatissimum L.). Theor. Appl. Genet. 119:53–63.CrossRefPubMedGoogle Scholar
  5. The Tomato Genome Consortium. 2012. The tomato genome sequence provides insights into fleshy fruit evolution. Nature 485:635–641.CrossRefGoogle Scholar
  6. Crisp, D. 1995. Radish, Raphanus sativus (Cruciferae), p.86–89 In: J. Smarttand N.W. Simmonds (eds.). Evolution of crop plants. Longman Scientific & Technical, Harlow, UK.Google Scholar
  7. De Candolle, A. 1886. Origin of cultivated plants. 2nd ed. Hafner Publishing Company, New York, NY, USA.Google Scholar
  8. Dieringer, D. and C. Schlotterer. 2003. MICROSATELLITEANALYSER (MSA): A platform independent analysis tool for large microsatellite data sets. Mol. Ecol. Notes 3:167–169.CrossRefGoogle Scholar
  9. Evanno, G., S. Regnaut, and J. Goudet. 2005. Detecting the number of clusters of individuals using the software STRUCTURE: A simulation study. Mol. Ecol. 14:2611–2620.CrossRefPubMedGoogle Scholar
  10. Hamilton J.P., S. Sim, K. Stoffel, A. Van Deynze, C.R. Buell, and D.M. Francis. 2012. Single nucleotide polymorphism discovery in cultivated tomato via sequencing by synthesis. Plant Genome-US 5:17–29.CrossRefGoogle Scholar
  11. Henslow, G. 1898. The history of the radish. Gard.Chron. 23:389.Google Scholar
  12. tHuang, S.W., R.Q. Li, Z.H. Zhang, L. Li, X.F. Gu, W. Fan, W.J. Lucas, X.W. Wang, B.Y. Xie, P.X. Ni, Y.Y. Ren, H.M. Zhu, J. Li, K. Lin, W.W. Jin, Z.J. Fei, G.C. Li, J. Staub, A. Kilian, E.a.G. Van Der Vossen, Y. Wu, J. Guo, J. He, Z.Q. Jia, Y. Ren, G. Tian, Y. Lu, J. Ruan, W.B. Qian, M.W. Wang, Q.F. Huang, B. Li, Z.L. Xuan, J.J. Cao, Asan, Z.G. Wu, J.B. Zhang, Q.L. Cai, Y.Q. Bai, B.W. Zhao, Y.H. Han, Y. Li, X.F. Li, S.H. Wang, Q.X. Shi, S.Q. Liu, W.K. Cho, J.Y. Kim, Y. Xu, K. Heller-Uszynska, H. Miao, Z.C. Cheng, S.P. Zhang, J. Wu, Y.H. Yang, H.X. Kang, M. Li, H.Q. Liang, X.L. Ren, Z.B. Shi, M. Wen, M. Jian, H.L. Yang, G.J. Zhang, Z.T. Yang, R. Chen, S.F. Liu, J.W. Li, L.J. Ma, H. Liu, Y. Zhou, J. Zhao, X.D. Fang, G.Q. Li, L. Fang, Y.R. Li, D.Y. Liu, H.K. Zheng, Y. Zhang, N. Qin, Z. Li, G.H. Yang, S. Yang, L. Bolund, K. Kristiansen, H.C. Zheng, S.C. Li, X.Q. Zhang, H.M. Yang, J. Wang, R.F. Sun, B.X. Zhang, S.Z. Jiang, J. Wang, Y.C. Du, and S.G. Li. 2009. The genome of the cucumber, Cucumis sativus L. Nat. Genet. 41:1275–1281.CrossRefGoogle Scholar
  13. Jarne, P. and P.J.L. Lagoda. 1996. Microsatellites, from molecules to populations and back. Trends Ecol. Evol. 11:424–429.CrossRefPubMedGoogle Scholar
  14. Jiang, L.N., L.J. Wang, L.W. Liu, X.W. Zhu, L.L. Zhai, and Y.Q. Cong. 2012. Development and characterization of cDNA library based novel EST-SSR marker in radish (Raphanus sativus L.). Sci. Hortic. 140:164–172.CrossRefGoogle Scholar
  15. Johnston, J.S., A.E. Pepper, A.E. Hall, Z.J. Chen, G. Hodnett, J. Drabek, R. Lopez, and H.J. Price. 2005. Evolution of genome size in Brassicaceae. Ann. Bot. 95:229–235.CrossRefPubMedCentralPubMedGoogle Scholar
  16. Kaneko, Y., C. Kimizuka-Takagi, S.W. Bang, and Y. Matsuzawa. 2007. Radish, p. 141–160. In:C. Kole (ed.).Genome mapping and molecular breeding in plants: Vegetables. Springer, Berlin Heidelberg, Germany.Google Scholar
  17. Kim, S., M. Park, S.I. Yeom, Y.M. Kim, J.M. Lee, H.A. Lee, E. Seo, J. Choi, K. Cheong, K.T. Kim, K. Jung, G.W. Lee, S.K. Oh, C. Bae, S.B. Kim, H.Y. Lee, S.Y. Kim, M.S. Kim, B.C. Kang, Y.D. Jo, H.B. Yang, H.J. Jeong, W.H. Kang, J.K. Kwon, C. Shin, J.Y. Lim, J.H. Park, J.H. Huh, J.S. Kim, B.D. Kim, O. Cohen, I. Paran, M.C. Suh, S.B. Lee, Y.K. Kim, Y. Shin, S.J. Noh, J. Park, Y.S. Seo, S.Y. Kwon, H.A. Kim, J.M. Park, H.J. Kim, S.B. Choi, P.W. Bosland, G. Reeves, S.H. Jo, B.W. Lee, H.T. Cho, H.S. Choi, M.S. Lee, Y. Yu, Y. Do Choi, B.S. Park, A. Van Deynze, H. Ashrafi, T. Hill, W.T. Kim, H.S. Pai, H.K. Ahn, I. Yeam, J.J. Giovannoni, J.K.C. Rose, I. Sorensen, S.J. Lee, R.W. Kim, I.Y. Choi, B.S. Choi, J.S. Lim, Y.H. Lee, and D. Choi. 2014. Genome sequence of the hot pepper provides insights into the evolution of pungency in Capsicum species. Nat. Genet. 46:270–278.CrossRefPubMedGoogle Scholar
  18. Kitamura, S. 1958. Varieties of radish and their transition, p. 1–19. In:I. Nishiyama(ed.). Japanese radish. The Japan Science Society Press, Tokyo, Japan.Google Scholar
  19. Kitashiba, H., F. Li, H. Hirakawa, T. Kawanabe, Z.W. Zou, Y. Hasegawa, K. Tonosaki, S. Shirasawa, A. Fukushima, S. Yokoi, Y. Takahata, T. Kakizaki, M. Ishida, S. Okamoto, K. Sakamoto, K. Shirasawa, S. Tabata, and T. Nishio. 2014. Draft sequences of the radish (Raphanus sativus L.) genome. DNA Res. 21:481–490.CrossRefPubMedCentralPubMedGoogle Scholar
  20. Kobabe, G. 1959. Naturliche Einkreuzung von Hederich (Raphanus raphanistrum L.) in Radies (Raphanus sativus var. radicula DC.) und das Verhalten von Knollenform und Farbe in den nachforgenden F-und R-Generationen. Zeitschrift Fur Pflanzenzuchtung 42:1–10.Google Scholar
  21. Koressaar, T. and M. Remm. 2007. Enhancements and modifications of primer design program Primer3. Bioinformatics 23:1289–1291.CrossRefPubMedGoogle Scholar
  22. Kwon, Y.S., S.G. Park, and S.I. Yi. 2009. Assessment of genetic variation among commercial tomato (Solanum lycopersicum L.) varieties using SSR markers and morphological characteristics. Genes Genom. 31:1–10.CrossRefGoogle Scholar
  23. Kwon, Y.S., Y.H. Oh, S.I. Yi, H.Y. Kim, J.M. An, S.G. Yang, S.H. Ok, and J.S. Shin. 2010. Informative SSR markers for commercial variety discrimination in watermelon (Citrullus lanatus). GenesGenom. 32:115–122.Google Scholar
  24. Nakatsuji, R., T. Hashida, N. Matsumoto, M. Tsuro, N. Kubo, and M. Hirai. 2011. Development of genomic and EST-SSR markers in radish (Raphanus sativus L.). Breed. Sci. 61:413–419.CrossRefPubMedCentralPubMedGoogle Scholar
  25. Nei, M. 1978. Estimation of average heterozygosity and genetic distance from a small number of individuals. Genetics 89:583–590.PubMedCentralPubMedGoogle Scholar
  26. Pritchard, J.K., M. Stephens, and P. Donnelly. 2000. Inference of population structure using multilocus genotype data. Genetics 155:945–959.PubMedCentralPubMedGoogle Scholar
  27. Ramchiary, N., V.D. Nguyen, X.N. Li, C.P. Hong, V. Dhandapani, S.R. Choi, G. Yu, Z.Y. Piao, and Y.P. Lim. 2011. Genic microsatellite markers in Brassica rapa: Development, characterization, mapping, and their utility in other cultivated and wild Brassica relatives. DNA Res. 18:305–320.CrossRefPubMedCentralPubMedGoogle Scholar
  28. Rohlf, F.J. 2008}. NTSYSpc: Numerical taxonomy system, ver.2.20. Exeter Publishing Ltd., Setauket, NY, USA.Google Scholar
  29. Shirasawa, K., M. Oyama, H. Hirakawa, S. Sato, S. Tabata, T. Fujioka, C. Kimizuka-Takagi, S. Sasamoto, A. Watanabe, M. Kato, Y. Kishida, M. Kohara, C. Takahashi, H. Tsuruoka, T. Wada, T. Sakai, and S. Isobe. 2011. An EST-SSR linkage map of Raphanus sativus and comparative genomics of the Brassicaceae. DNA Res. 18: 221–232.CrossRefPubMedCentralPubMedGoogle Scholar
  30. Sim, S., G. Durstewitz, J. Plieske, R. Wieseke, M.W. Ganal, A. Van Deynze, J.P. Hamilton, C.R. Buell, M. Causse, S. Wijeratne, and D.M. Francis. 2012. Development of a large SNP genotyping array and generation of high-density genetic maps in tomato. PLoS ONE 7:e40563.CrossRefGoogle Scholar
  31. Sim, S., J.K. Yu, Y.K. Jo, M.E. Sorrells, and G. Jung. 2009. Transferability of cereal EST-SSR markers to ryegrass. Genome 52:431–437.CrossRefPubMedGoogle Scholar
  32. Varshney, R.K., A. Graner, and M.E. Sorrells. 2005. Genic microsatellite markers in plants: Features and applications. Trends Biotechnol. 23:48–55.CrossRefPubMedGoogle Scholar
  33. Wang, S.F., X.F. Wang, Q.W. He, X.X. Liu, W.L. Xu, L.B. Li, J.W. Gao, and F.D. Wang. 2012. Transcriptome analysis of the roots at early and late seedling stages using Illumina paired-end sequencing and development of EST-SSR markers in radish. Plant Cell Rep. 31:1437–1447.CrossRefPubMedGoogle Scholar
  34. Wang, X.W., H.Z. Wang, J. Wang, R.F. Sun, J. Wu, S.Y. Liu, Y.Q. Bai, J.H. Mun, I. Bancroft, F. Cheng, S.W. Huang, X.X. Li, W. Hua, J.Y. Wang, X.Y. Wang, M. Freeling, J.C. Pires, A.H. Paterson, B. Chalhoub, B. Wang, A. Hayward, A.G. Sharpe, B.S. Park, B. Weisshaar, B.H. Liu, B. Li, B. Liu, C.B. Tong, C. Song, C. Duran, C.F. Peng, C.Y. Geng, C.S. Koh, C.Y. Lin, D. Edwards, D.S. Mu, D. Shen, E. Soumpourou, F. Li, F. Fraser, G. Conant, G. Lassalle, G.J. King, G. Bonnema, H.B. Tang, H.P. Wang, H. Belcram, H.L. Zhou, H. Hirakawa, H. Abe, H. Guo, H. Wang, H.Z. Jin, I.A.P. Parkin, J. Batley, J.S. Kim, J. Just, J.W. Li, J.H. Xu, J. Deng, J.A. Kim, J.P. Li, J.Y. Yu, J.L. Meng, J.P. Wang, J.M. Min, J. Poulain, J. Wang, K. Hatakeyama, K. Wu, L. Wang, L. Fang, M. Trick, M.G. Links, M.X. Zhao, M.N. Jin, N. Ramchiary, N. Drou, P.J. Berkman, Q.L. Cai, Q.F. Huang, R.Q. Li, S. Tabata, S.F. Cheng, S. Zhang, S.J. Zhang, S.M. Huang, S. Sato, S.L. Sun, S.J. Kwon, S.R. Choi, T.H. Lee, W. Fan, X. Zhao, X. Tan, X. Xu, Y. Wang, Y. Qiu, Y. Yin, Y.R. Li, Y.C. Du, Y.C. Liao, Y. Lim, Y. Narusaka, Y.P. Wang, Z.Y. Wang, Z.Y. Li, Z.W. Wang, Z.Y. Xiong, and Z.H. Zhang. 2011. The genome of the mesopolyploid crop species Brassica rapa. Nat. Genet. 43:1035–1039.CrossRefPubMedGoogle Scholar
  35. Weir, B.S. and C.C. Cockerham. 1984. Estimating F-statistics for the analysis of population-structure. Evolution 38:1358–1370.CrossRefGoogle Scholar
  36. Yagi, M., T. Kimura, T. Yamamoto, S. Isobe, S. Tabata, and T. Onozaki. 2012. QTL analysis for resistance to bacterial wilt (Burkholderia caryophylli) in carnation (Dianthus caryophyllus) using an SSR-based genetic linkage map. Mol. Breed. 30:495–509.CrossRefGoogle Scholar
  37. Yamagishi, H. 2004. Assessment of cytoplasmic polymorphisms by PCR-RFLP of the mitochondrial orfB region in wild and cultivated radishes. Plant Breed. 123:141–144.CrossRefGoogle Scholar
  38. Yamagishi, H. and T. Terachi. 2003. Multiple origins of cultivated radishes as evidenced by a comparison of the structural variations in mitochondrial DNA of Raphanus. Genome 46:89–94.CrossRefPubMedGoogle Scholar
  39. Yamane, K., N. Lu, and O. Ohnishi. 2005. Chloroplast DNA variations of cultivated radish and its wild relatives. Plant Sci. 168:627–634.CrossRefGoogle Scholar
  40. Yamane, K., N. Lu, and O. Ohnishi. 2009. Multiple origins and high genetic diversity of cultivated radish inferred from polymorphism in chloroplast simple sequence repeats. Breed. Sci. 59:55–65.CrossRefGoogle Scholar
  41. Yi, S.I., K.M. Bae, Y.S. Kwon, and I.H. Cho. 2006. Development of oriental melon (Cucumis melo L.)derived SSR markers using a PCR-based method and polymorphic application for the genotyping of commercial lines. Kor. J. Genet. 28:317–324.Google Scholar

Copyright information

© Korean Society for Horticultural Science and Springer-Verlag GmbH 2015

Authors and Affiliations

  • Kyung-Mi Bae
    • 1
  • Sung-Chur Sim
    • 2
  • Jee-Hwa Hong
    • 3
  • Keun-Jin Choi
    • 4
  • Do-Hoon Kim
    • 5
  • Yong-Sham Kwon
    • 5
    Email author
  1. 1.Department of IT-Agriculture EcologyKorea Vocational College of Food & AgricultureSeoulKorea
  2. 2.Department of Bioresources EngineeringSejong UniversitySeoulKorea
  3. 3.Seed Testing & Research CenterKorea Seed & Variety Service, Ministry for Agriculture, Food and Rural AffairsGimcheonKorea
  4. 4.Vegetable Research Division, National Institute of Horticultural & Herbal ScienceRural Development AdministrationWanjuKorea
  5. 5.Department of Genetic EngineeringCollege of Natural Resources and Life Science, Dong-A UniversityBusanKorea

Personalised recommendations