Hostname: page-component-78c5997874-ndw9j Total loading time: 0 Render date: 2024-11-07T01:41:54.974Z Has data issue: false hasContentIssue false
  • English
  • Français

Evaluation of PCR methods for fixed bivalve larvae

Published online by Cambridge University Press:  08 July 2008

Hideki Sawada ,
Hajime Saito ,
Masatomi Hosoi  and
Haruhiko Toyohara
Hideki Sawada
Affiliation:
Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan
Hajime Saito
Affiliation:
National Research Institute of Fisheries Engineering, Fisheries Research Agency, Hasaki, Kamisu, Ibaraki 314-0408, Japan
Masatomi Hosoi
Affiliation:
Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan Present address: School of Environmental Science, University of Shiga Prefecture, 2500 Hassaka-cho, Hikone-City, Shiga 522-8533, Japan
Haruhiko Toyohara*
Affiliation:
Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan
*
Correspondence should be addressed to: Haruhiko Toyohara, Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan email: [email protected]
Get access Rights & Permissions [Opens in a new window]

Abstract

Investigating the spatio-temporal patterns of planktonic larvae is fundamental to studies regarding stock assessment and larval dispersal of commercial and non-commercial, i.e. invasive or rare marine invertebrates. Because of the difficulty involved in morphological identification of marine invertebrate larvae, various molecular methods based on PCR have been used to enhance taxonomic resolution. In previous studies, different methods for the preservation or pretreatment of larvae were applied in each case. However, no comparative studies have been conducted to determine the optimal method for PCR testing for bivalve larvae, and no information is available regarding the selection of an appropriate method.

This study compared the PCR success rate of 6 pretreatment methods for larvae of the Mediterranean blue mussel, which was preserved using different fixatives (70% ethanol, 100% ethanol, 70% acetone and 10% formalin). The results revealed that the success rate of PCR was different for each pretreatment; moreover, the use of ammonium sulphate and Tween 20 buffer with proteinase K digestion was found to be the most effective method. Some pretreatments showed lower success rates for long-fixed larvae than for short-fixed larvae for formalin-fixed larvae; however, the success rate of PCR amplification for ethanol-fixed larvae pretreated by this method did not decrease through 1-year fixation. In addition, this pretreatment showed a high success rate for different fixation periods. These findings suggest that the selection of the pretreatment method is critically important for successfully amplifying larval DNA and that the pretreatment involving the use of ammonium sulphate prior to PCR amplification enables the use of fixatives for preserving bivalve larvae. This method will be utilized in various field studies and molecular genetic studies.

Keywords

fixativeDNA extractionbivalvelarvaeidentificationdispersalPCR
Type
Research Article
Information
Journal of the Marine Biological Association of the United Kingdom , Volume 88 , Issue 7 , November 2008 , pp. 1441 - 1449
Copyright
Copyright © Marine Biological Association of the United Kingdom 2008

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Abu Al-Soud, W. and Radstrom, P. (2000) Effects of amplification facilitators on diagnostic PCR in the presence of blood, feces, and meat. Journal of Clinical Microbiology 38, 4463–4470.CrossRefGoogle ScholarPubMed
Andre, C., Lindegarth, M., Jonsson, P.R. and Sundberg, P. (1999) Species identification of bivalve larvae using random amplified polymorphic DNA (RAPD): differentiation between Cerastoderma edule and C. lamarcki. Journal of the Marine Biological Association of the United Kingdom 79, 563–565.CrossRefGoogle Scholar
Baldwin, B.S., Black, M., Sanjur, O., Gustafson, R., Lutz, R.A. and Vrijenhoek, R.C. (1996) A diagnostic molecular marker for zebra mussels (Dreissena polymorpha) and potentially co-occurring bivalves: mitochondrial COI. Molecular Marine Biology and Biotechnology 5, 9–14.Google ScholarPubMed
Bancroft, J.D. and Stevens, A. (2002) Theory and practice of histological techniques. New York: Churchill Livingstone.Google Scholar
Beaumont, A.R. and Abdulmatin, A.K.M. (1994) Differences in morphology, survival and size between self-fertilized and cross-fertilized larvae of Mytilus galloprovincialis. Journal of the Marine Biological Association of the United Kingdom 74, 445–448.CrossRefGoogle Scholar
Bell, J.L. and Grassle, J.P. (1998) A DNA probe for identification of larvae of the commercial surfclam (Spisula solidissima). Molecular Marine Biology and Biotechnology 7, 127–137.Google ScholarPubMed
Bendezu, I.F., Slater, J.W. and Carney, B.F. (2005) Identification of Mytilus spp. and Pecten maximus in Irish waters by standard PCR of the 18S rDNA gene and multiplex PCR of the 16S rDNA gene. Marine Biotechnology 7, 687–696.CrossRefGoogle ScholarPubMed
Bierne, N., Launey, S., Naciri-Graven, Y. and Bonhomme, F. (1998) Early effect of inbreeding as revealed by microsatellite analyses on Ostrea edulis larvae. Genetics 148, 1893–1906.CrossRefGoogle ScholarPubMed
Bucklin, A. (2000) Methods for population genetic analysis of zooplankton. In Harris, R., Wiebe, P., Lenz, J., Skjoldal, H-R. and Huntley, M. (eds) ICES Zooplankton Methodology Manual. London: Academic Press, pp. 533–570.CrossRefGoogle Scholar
Burzynski, A., Zbawicka, M., Skibinski, D.O.F. and Wenne, R. (2003) Evidence for recombination of mtDNA in the marine mussel Mytilus trossulus from the Baltic. Molecular Biology and Evolution 20, 388–392.CrossRefGoogle ScholarPubMed
Côrte-Real, H., Holland, P.W.H. and Dixon, D.R. (1994) Inheritance of a nuclear-DNA polymorphism assayed in single bivalve larvae. Marine Biology 120, 415–420.CrossRefGoogle Scholar
Dawson, M.N., Raskoff, K.A. and Jacobs, D.K. (1998) Field preservation of marine invertebrate tissue for DNA analyses. Molecular Marine Biology and Biotechnology 7, 145–152.Google ScholarPubMed
Deagle, B.E., Bax, N. and Patil, J.G. (2003) Development and evaluation of a PCR-based test for detection of Asterias (Echinodermata: Asteroidea) larvae in Australian plankton samples from ballast water. Marine and Freshwater Research 54, 709–719.CrossRefGoogle Scholar
Dessauer, H.C., Cole, C.J. and Hafner, M.S. (1996) Collection and storage of tissues. In Hillis, D.M. and Moritz, C. (eds) Molecular systematics. Sunderland, MA: Sinauer Associates, pp. 29–41.Google Scholar
Evans, B.S., White, R.W.G. and Ward, R.D. (1998) Genetic identification of asteroid larvae from Tasmania, Australia, by PCR-RFLP. Molecular Ecology 7, 1077–1082.CrossRefGoogle Scholar
Frischer, M.E., Danforth, J.M., Tyner, L.C., Leverone, J.R., Marelli, D.C., Arnold, W.S. and Blake, N.J. (2000) Development of an Argopecten-specific 18S rRNA targeted genetic probe. Marine Biotechnology 2, 11–20.CrossRefGoogle ScholarPubMed
Frischer, M.E., Hansen, A.S., Wyllie, J.A., Wimbush, J., Murray, J. and Nierzwick-Bauer, S.A. (2002) Specific amplification of the S-18 rRNA gene as a method to detect zebra mussel (Dreissena polymorpha) larvae in plankton samples. Hydrobiologia 487, 33–44.CrossRefGoogle Scholar
Fukatsu, T. (1999) Acetone preservation: a practical technique for molecular analysis. Molecular Ecology 8, 1935–1945.CrossRefGoogle ScholarPubMed
Griffin, H.G. and Griffin, A.M. (1994) PCR technology current innovations. Boca Raton, FL, CRC Press.CrossRefGoogle Scholar
Hare, M.P., Palumbi, S.R. and Butman, C.A. (2000) Single-step species identification of bivalve larvae using multiplex polymerase chain reaction. Marine Biology 137, 953–961.CrossRefGoogle Scholar
Hosoi, M., Hosoi-Tanabe, S., Sawada, H., Ueno, M., Toyohara, H. and Hayashi, I. (2004) Sequence and polymerase chain reaction–restriction fragment length polymorphism analysis of the large subunit rRNA gene of bivalve: simple and widely applicable technique for multiple species identification of bivalve larva. Fisheries Science 70, 629–637.CrossRefGoogle Scholar
Ikegami, S., Mitsuno, T., Kataoka, M., Yajima, S. and Komatsu, M. (1991) Immunological survey of planktonic embryos and larvae of the starfish Asterina pectinifera, obtained from the sea, using a monoclonal-antibody directed against egg polypeptides. Biological Bulletin. Marine Biological Laboratory, Woods Hole 181, 95–103.CrossRefGoogle ScholarPubMed
Inadome, Y. and Noguchi, M. (2003) Selection of higher molecular weight genomic DNA for molecular diagnosis from formalin-fixed material. Diagnostic Molecular Pathology 12, 231–236.CrossRefGoogle ScholarPubMed
Inoue, K., Waite, J.H., Matsuoka, M., Odo, S. and Harayama, S. (1995) Interspecific variations in adhesive protein sequences of Mytilus edulis, M. galloprovincialis, and M. trossulus. Biological Bulletin. Marine Biological Laboratory, Woods Hole 189, 370–375.CrossRefGoogle ScholarPubMed
Kasyanov, V.L. (1984) Planktotrophic larvae of bivalve mollusks: morphology, physiology and behaviour. Biol Morya, Kiev 10, 117–128.Google Scholar
Larsen, J.B., Frischer, M.E., Rasmussen, L.J. and Hansen, B.W. (2005) Single-step nested multiplex PCR to differentiate between various bivalve larvae. Marine Biology 146, 1119–1129.CrossRefGoogle Scholar
Launey, S. and Hedgecock, D. (2001) High genetic load in the Pacific oyster Crassostrea gigas. Genetics 159, 255–265.CrossRefGoogle ScholarPubMed
Lewin, R. (1986) Supply-side ecology. Science 234, 25–27.CrossRefGoogle ScholarPubMed
Livi, S., Cordisco, C., Damiani, C., Romanelli, M. and Crosetti, D. (2006) Identification of bivalve species at an early developmental stage through PCR-SSCP and sequence analysis of partial 18S rDNA. Marine Biology 149, 1149–1161.CrossRefGoogle Scholar
Lopez-Pinon, M.J., Insua, A. and Mendez, J. (2002) Identification of four scallop species using PCR and restriction analysis of the ribosomal DNA internal transcribed spacer region. Marine Biotechnology 4, 495–502.CrossRefGoogle ScholarPubMed
McEdward, L. (1995) Ecology of marine invertebrate larvae. Boca Raton, FL: CRC Press.Google Scholar
Medeiros-Bergen, D.E., Olson, R.R., Conroy, J.A. and Kocher, T.D. (1995) Distribution of holothurian larvae determined with species-specific genetic probes. Limnology and Oceanography 40, 1225–1235.CrossRefGoogle Scholar
Morgan, T.S. and Rogers, A.D. (2001) Specificity and sensitivity of microsatellite markers for the identification of larvae. Marine Biology 139, 967–973.Google Scholar
Olive, D.M., Simsek, M. and Al-Mufti, S. (1989) Polymerase chain reaction assay for detection of human cytomegalovirus. Journal of Clinical Microbiology 27, 1238–1242.CrossRefGoogle ScholarPubMed
Olson, R.R., Runstadler, J.A. and Kocher, T.D. (1991) Whose larvae? Nature 351, 357–358.CrossRefGoogle ScholarPubMed
Passamonti, M., Boore, J.L. and Scali, V. (2003) Molecular evolution and recombination in gender-associated mitochondrial DNAs of the manila clam Tapes philippinarum. Genetics 164, 603–611.CrossRefGoogle ScholarPubMed
Patil, J.G., Gunasekera, R.M., Deagle, B.E. and Bax, N.J. (2005) Specific detection of Pacific oyster (Crassostrea gigas) larvae in plankton samples using nested polymerase chain reaction. Marine Biotechnology 7, 11–20.CrossRefGoogle ScholarPubMed
Paugam, A., Le Pennec, M., Marhic, A. and Andre-Fontaine, G. (2003) Immunological in situ determination of Pecten maximus larvae and their temporal distribution. Journal of the Marine Biological Association of the United Kingdom 83, 1083–1093.CrossRefGoogle Scholar
Rolfs, A. (1992) PCR: clinical diagnostics and research. Berlin: Springer-Verlag.CrossRefGoogle Scholar
Sambrook, J. and Russell, D.W. (2001) Molecular cloning: a laboratory manual. New York: Cold Spring Harbor Laboratory Press.Google Scholar
Shi, S.R., Cote, R.J., Wu, L., Liu, C., Datar, R., Shi, Y., Liu, D., Lim, H. and Taylor, C.R. (2002) DNA extraction from archival formalin-fixed, paraffin-embedded tissue sections based on the antigen retrieval principle: heating under the influence of pH. Journal of Histochemistry and Cytochemistry 50, 1005–1011.CrossRefGoogle ScholarPubMed
Shi, S.R., Datar, R., Liu, C., Wu, L., Zhang, Z., Cote, R.J. and Taylor, C.R. (2004) DNA extraction from archival formalin-fixed, paraffin-embedded tissues: heat-induced retrieval in alkaline solution. Histochemistry and Cell Biology 122, 211–218.CrossRefGoogle ScholarPubMed
Sokal, R.R. and Rohlf, F.J. (1994) Biometry. New York: W.H. Freeman and Co.Google Scholar
Srinivasan, M., Sedmak, D. and Jewell, S. (2002) Effect of fixatives and tissue processing on the content and integrity of nucleic acids. American Journal of Pathology 161, 1961–1971.CrossRefGoogle ScholarPubMed
Sutherland, B., Stewart, D., Kenchington, E.R. and Zouros, E. (1998) The fate of paternal mitochondrial DNA in developing female mussels, Mytilus edulis: implications for the mechanism of doubly uniparental inheritance of mitochondrial DNA. Genetics 148, 341–347.CrossRefGoogle ScholarPubMed
Taris, N., Baron, S., Sharbel, T.F., Sauvage, C. and Boudry, P. (2005) A combined microsatellite multiplexing and boiling DNA extraction method for high-throughput parentage analyses in the Pacific oyster (Crassostrea gigas). Aquaculture Research 36, 516–518.CrossRefGoogle Scholar
Toro, J.E. (1998) Molecular identification of four species of mussels from southern Chile by PCR-based nuclear markers: the potential use in studies involving planktonic surveys. Journal of Shellfish Research 17, 1203–1205.Google Scholar
Toro, J.E., Innes, D.J. and Thompson, R.J. (2004) Genetic variation among life-history stages of mussels in a Mytilus edulis–M. trossulus hybrid zone. Marine Biology 145, 713–725.Google Scholar
Wilson, I.G. (1997) Inhibition and facilitation of nucleic acid amplification. Applied and Environmental Biology 63, 3741–3751.CrossRefGoogle ScholarPubMed
Wood, A.R., Beaumont, A.R., Skibinski, D.O.F. and Turner, G. (2003) Analysis of a nuclear-DNA marker for species identification of adults and larvae in the Mytilus edulis complex. Journal of Molluscan Studies 69, 61–66.CrossRefGoogle Scholar