Ocean Science Journal

, Volume 40, Issue 3, pp 43–54 | Cite as

Molecular analysis of complete ssu to lsu rdna sequence in the harmful dinoflagellatealexandrium tamarense (korean isolate, HY970328M)



New PCR primers (N=18) were designed for the isolation of complete SSU to LSU rDNA sequences from the dinoflagellateAlexandrium tamarense. Standard PCR, employing each primer set selected for amplifications of less than 1.5 kb, successfully amplified the expected rDNA regions of A. tamarense (Korean isolate, HY970328M). Complete SSU, LSU rDNAs and ITS sequences, including 5.8S rDNA, were recorded at 1,800 bp, 520 bp and 3,393 bp, respectively. The LSU rDNA sequence was the first report inAlexandrium genus. No intron was found in the LSU rRNA coding region. Twelve D-domains within the LSU rDNA were put together into 1,879 bp (44.4% G+C), and cores into 1514 bp (42.8% G+C). The core sequence was significantly different (0.0867 of genetic distance, 91% sequence similarity) in comparison withProrocentrum micans (GenBank access. no. X16108). The D2 region was the longest in length (300 bp) and highly variable among the 12 D-domains. In a phylogenetic analysis using complete LSU rDNA sequences of a variety of phytoplankton,A tamarense was clearly separated with high resolution against other species. The result suggests that the sequence may resolve the taxonomic ambiguities ofAlexandrium genus, particularly of the tamarensis complex.

Key words

Alexandrium tamarense large-subunit rDNA phylogenetic analysis D-domain 


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  1. Adachi, M., Y. Sako, and Y. Ishida. 1994. Restriction fragment length polymorphism of ribosomal DNA internal transcribed spacer and 5.8S regions in JapaneseAlexandrium species (Dinophyceae).J. Phycol. 30, 857–863.CrossRefGoogle Scholar
  2. Adachi, M., Y. Sako, and Y. Ishida. 1996. Analysis ofAlexandrium (Dinophyceae) species using sequences of the 5.8S ribosomal DNA and internal transcribed spacer regions.J. Phycol. 32, 424–32.CrossRefGoogle Scholar
  3. Anderson, D.M. 1989. Cysts as factors inPyrodinium bahamense ecology. In: Hallegraeff, G.M., and J.L. Maclean (eds.), Biology, Epidemiology and Management ofPyrodinium Red Tides. ICLARM Conference Proceedings21, 81–88.Google Scholar
  4. Anderson, D.M., D.M. Kulis, G.J. Doucette, J.C. Gallagher, and E. Balech. 1994. Biogeography of toxic dinoflagellates in the genus Alexandrium from the northeastern United States and Canada.Mar. Biol. 120, 467–478.CrossRefGoogle Scholar
  5. Ben Ali, A., J. Wuyts, R. De Wachter, A. Meyer, Y. Van de Peer. 1999. Construction of a variability map for eukaryotic large subunit ribosomal RNA.Nucleic Acids Res. 27(14), 2825–2831.CrossRefGoogle Scholar
  6. Ben Ali A., R. De Baere, G. Van der Auwera, R. De Wachter, Y. Van de Peer. 2001. Phylogenetic relationships among algae based on complete large-subunit rRNA sequences.Int. J. Syst. Evol. Microbiol. 51, 737–749.Google Scholar
  7. Cembella, A.D., J.J. Sullivan, G.L. Boyer, F.J.R. Taylor, and R.J. Anderson. 1987. Variations in paraly tic shellfish toxin composition within theProtogonyaulax tamarensis/catenella species complex: red tide dinoflagelllates.Biochem. Syst. Ecol. 15, 171–186.CrossRefGoogle Scholar
  8. Cembella, A.D., F.J.R. Taylor, and J.C. Therriault. 1988. Cladistic analysis of electrophorectic variants within the toxic dinoflagellate genusProtogonyaulax.Bot. Mar. 31, 39–51.CrossRefGoogle Scholar
  9. Destombe, C., A.D. Cembella, C.A. Murphy, and M.A. Ragan. 1992. Nucleotide sequence of the18S ribosomal RNA genes from the marine dinoflagellateAlexandrium tamarense (Gonyalulacales, Dinophyta).Phycologia 31, 121–124.Google Scholar
  10. Guillard, R.R.L., and J.H. Ryther. 1962. Studies of marine planktonic diatoms 1.Cyclotella nana Hustedt, andDetonula confervacea (Cleve) Gran.Can. J. Microbio. 8, 229–39.Google Scholar
  11. Hayhome, B.A., D.M. Anderson, D.M. Kulis, and D.J. Whitten. 1989. Variation among congeneric dinofalgellates from the northeastern United States and Canada. I. Enzyme electrophoresis.Mar. Biol. 101, 427–435.CrossRefGoogle Scholar
  12. Higman, W.A., D.M., Stone, and J.M. Lewis. 2001. Sequence comparisons of toxic and non-toxicAlexandrium tamarense (Dinophyceae) isolates from UK waters.Phylologia 40, 256- 262.Google Scholar
  13. Hirashita, T., K. Ichimi, S. Montani, M. Nomura, and S. Tajima. 2000. Molecular analysis of ribosomal RNA gene of red tide algae obtained from the Seto Inland Sea.Mar. Biotechnol. 2, 267–273.Google Scholar
  14. Ki, J.-S, and Han M.-S. 2005. Efficient 5′ ETS walking from conserved 18S rDNA sequences of dinoflagellatesAlexandrium andAkashiwo sanguinea (Dinophyceae).J. Appl. Phycol. (In press).Google Scholar
  15. Ki, J.-S., G. Y. Jang, and M.-S. Han. 2004. Integrated method for single-cell DNA extraction, PCR amplification, and sequencing of the ribosomal DNA from the harmful dinoflagellatesCochlodinium polykrikoides andAlexandrium catenella.Mar. Biotechnol. 6, 587–593.CrossRefGoogle Scholar
  16. Kim, C.-J., Y. Sako, A. Uchida, and C.-H. Kim. 2004. Molecular phylogenetic relationships within the genusAlexandrium (Dinophyceae) based on the nuclear-encoded SSU and LSU rDNA D1-D2 sequences.J. Kor. Soc. Oceanogr. 39, 172–185.Google Scholar
  17. Kim, K.-Y., and C.-H. Kim. 2004. A molecular phylogenetic study on KoreanAlexandrium catenella andA. tamarense isolates (Dinophyceae) based on the partial LSU rDNA sequence data.J. Kor. Soc. Oceanogr. 39, 163–171.Google Scholar
  18. Kimura, M. 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences.J. Mol. Evol. 16, 111–120.CrossRefGoogle Scholar
  19. Kumar, S. K. Tamura, M. and Nei. 2004. MEGA3: Integrated Software for Molecular Evolutionary Genetics Analysis and Sequence Alignment Briefings in Bioinformatics5, 150–163.Google Scholar
  20. Lenaers, G., L. Maroteau, B. Michot, and M. Herzog. 1998. Dinoflagellates in evolution: a molecular phylogenetic analysis of large subunit ribosomal RNA.J. Mol. Evol. 29, 40–51.CrossRefGoogle Scholar
  21. Medlin, L., H.J. Elwood, S. Stickel, and M.L. Sogin. 1988. The characterization of enzymatically amplifed eukaryotic 16S- like rRNA-coding regions.Gene 71, 491–499.CrossRefGoogle Scholar
  22. Rae, P.M.M. 1970. The nature and processing of ribosomal nucleic acid in a dinoflagellate.J. Cell Biol. 46, 106–113.CrossRefGoogle Scholar
  23. Sako, Y. 2000. Phylogenetic analysis and molecular identification of harmful dinoflagellates.Bull. Plankton Soc. Japan 47, 46–55.Google Scholar
  24. Sako, Y., M. Adachi, and Y. Ishida. 1993. Preparation and characterization of monoclonal antibodies toAlexandrium sepcies. In: Smayda, T.J. and Z. Shimizu. (eds) Toxic Phytoplankton Blooms in the Sea. Elsevier, New York, pp. 87–93.Google Scholar
  25. Sako, Y. , C.H. Kim, H. Ninomiya, M. Adachi, and Y. Ishida. 1990. Isozyme and cross analysis of mating populations in theAlexandrium catenella/ tamarense species complex. In: Granili, E., B. Sundstrom, L. Eldler, and D.M. Anderson (ededs) Toxic Phytoplankton Blooms in the Sea. Elsevier, pp. 95-102.Google Scholar
  26. Sambrook, J., E.F. Fritsch, and T. Maniatis. 1989. Molecular cloning: a laboratory manual. Cold Spring Harbor Press, New York, NY.Google Scholar
  27. Scholin, C.A., and D.M. Anderson. 1996. LSU rDNA-based assays for discriminating species and strains ofAlexandrium (Dinophyceae).J. Phycol. 32, 1022–1035.CrossRefGoogle Scholar
  28. Scholin, C.A., M. Herzog, M.L. Sogin, and D.M. Anderson. 1994. Identification of group and strain-specific genetic markers for globally distributedAlexandrium (Dinophyceae). Part II. Sequence analysis of a fragment of the LSU rRNA gene.J. Phycol. 30, 999–1011.CrossRefGoogle Scholar
  29. Usup, G., L.C. Pin, A.A Ahmad, and L.P. Teen. 2002. Phylogeneitc realtionship ofAlexandrium tamiyavanichii (Dynophyceae) to otherAlexandrium speices based on ribosomal RNA gene sequences.Harmful Algae. 1, 59–68.CrossRefGoogle Scholar

Copyright information

© Korea Ocean Research and Development Institute(KORDI) and the Korean Society of Oceanography(KSO) 2005

Authors and Affiliations

  1. 1.Department of Life ScienceCollege of Natural Sciences, Hanyang UniversitySeoulKorea

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