Molecular Breeding

, Volume 13, Issue 3, pp 263–279 | Cite as

Efficient screening for expressed sequence tag polymorphisms (ESTPs) by DNA pool sequencing and denaturing gradient gel electrophoresis (DGGE) in spruces

Article

Abstract

There is an urgent need to accelerate the development of informative codominant markers of coding regions such as ESTPs (expressed sequence tag polymorphisms) to estimate map synteny within and among taxa. A set of primer pairs for 207 ESTs or cDNAs from Picea and Pinus taxa was screened on three distantly-related taxa in the genus Picea, P. mariana (Mill.) B.S.P., P. glauca (Moench) Voss and P. abies (L.) Karst. Of these, 118 (57%) resulted in positive amplification of single-locus gene products in the first two species. To detect polymorphism, these 118 markers were further screened on a panel of 10 pedigree parents for each of P. mariana and P. glauca, either by agarose gel electrophoresis (AGE) or by parallel denaturing gradient gel electrophoresis (DGGE) with standard conditions of 15-45% urea-formamide. Of these, 87 and 74 were found polymorphic in P. mariana and P. glauca, respectively, and 65 were polymorphic in both species. DNA pool sequencing has been explored as a possible strategy to increase economically the detection throughput of SNPs and small indels, and to characterize the types of DNA polymorphism detected by DGGE. Different DNA samples of known sequences were pooled in different ratio mixtures before and after PCR amplifications to determine their minimum relative abundance for detection of DNA polymorphisms by sequencing. For detection of a polymorphism in the DNA pools, the minimum level of relative abundance was 10%. Pooling DNA samples before or after PCR amplification had no effect on the detection of polymorphism by sequencing. For each species panel, the DNAs were pooled and then amplified and sequenced for the 118 primer pairs. With this strategy, the number of ESTPs increased to 107 in P. mariana and 106 in P. glauca, and the number of ESTPs shared by both species increased to 99. About half of the ESTP markers displayed both SNP and indel polymorphisms while the other half displayed only SNPs. Most of the additional ESTPs were amenable to detection by DGGE or CAPS (Cleaved Amplified Polymorphic Sequence) for mapping purposes.

Codominant markers Conifers Consensus mapping Insertion-deletion Picea Single nucleotide polymorphism 

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References

  1. Ahmadian A. and Lundeberg J. 2002. A brief history of geneticvariation analysis. Biotechniques 32: 1122–1137.Google Scholar
  2. Babula D., Kaczmarek M., Barakat A., Delseny M., Quiros C.F. and Sadowski J. 2003. Chromosomal mapping of Brassica ol-eraceabased on ESTs from Arabidopsis thaliana: complexity ofthe comparative map. Mol. Genet. Genomics 268: 656–665.Google Scholar
  3. Barcellos L.F., Klitz W., Field L.L., Tobias R., Bowcock A.M., Wilson R., Nelson M.P., Nagatomi J. and Thomson G. 1997. Association mapping of disease loci, by use of a pooled DNA genomic screen. Am. J. Hum. Genet. 61: 734–747.Google Scholar
  4. Bhattramakki D., Dolan M., Hanafey M., Wineland R., Vaske D., Register J.C., Tingey S.V. and Rafalski A. 2002. Insertion-deletion polymorphisms in 3'regions of maize genes occur frequently and can be used as highly informative genetic markers. Plant Mol. Biol. v48: 539–547.Google Scholar
  5. Brown G.R., Kadel E.E., Bassoni D.L., Kiehne K.L., Temesgen B., van Buijtenen J.P., Sewell M.M., Marshall K.A. and Neale D.B. 2001. Anchored reference loci in loblolly pine Pinus taeda L.for integrating pine genomics. Genetics 159: 799–809.Google Scholar
  6. Cargill M., Altshuler D., Ireland J., Sklar P., Ardlie K., Patil N., Shaw N., Lane C.R., Lim E.P., Kalyanaraman N., Nemesh J., Ziaugra L., Friedland L., Rolfe A., Warrington J., Lipshutz R., Daley G.Q. and Lander E.S. 1999. Characterization of single-nucleotide polymorphisms in coding regions of human genes. Nature Genet. 22: 231–238.Google Scholar
  7. Ching A., Caldwell K.S., Jung M., Dolan M., Smith O.S., TingeyS., Morgante M. and Rafalski A.J. 2002. SNP frequency, haplo-type structure and linkage disequilibrium in elite maize inbred lines. BMC Genet. 3: 19–33.Google Scholar
  8. Choy Y.S., Dabora S.L., Hall F., Ramesh V., Niida Y., Franz D., Kasprzyk-Obara J., Reeve M.P. and Kwiatkowski D.J. 1999. Superiority of denaturing high performance liquid chromatography over single-stranded conformation and conformation-sensitive gel electrophoresis for mutation detection in TSC2. Ann. Hum. Genet. v63: 383–391.Google Scholar
  9. Fournier D., Perry D.J., Beaulieu J., Bousquet J. and Isabel N. 2002. Optimizing expressed sequence TAG polymorphisms by single strand conformation polymorphism in spruces. For. Genet. 9: 11–17.Google Scholar
  10. Gamache I., Jaramillo-Corea J.P., Payette S. and Bousquet J. 2003. Diverging patterns of mitochondrial and nuclear DNA diversity in sub arctic black spruce: imprint of a founder effect associated with postglacial colonization. Mol. Ecol. 2: 891–901.Google Scholar
  11. Germer S., Holland M.J. and Higuchi R. 2000. High-throughputSNP allele-frequency determination in pooled DNA samples bykinetic PCR. Genome Res. v10: 258–266.Google Scholar
  12. Goddard K.A., Hopkins P.J., Hall J.M. and Witte J.S. 2000. Linkage disequilibrium and allele-frequency distributions for 114 single-nucleotide polymorphisms in five populations. Am. J. Hum. Genet. 66: 216–234.Google Scholar
  13. Gosselin I., Zhou Y., Bousquet J. and Isabel N. 2002. Megagame-tophyte-derived linkage maps of white spruce Picea glauca based on RAPD, SCAR and ESTP markers. Theor. Appl. Genet. 104: 987–997.Google Scholar
  14. Grivet L., Glaszmann J.C., Vincentz M., da Silva F. and Arruda P. 2003. ESTs as a source for sequence polymorphism discovery in sugar cane: example of the Adh genes. Theor. Appl. Genet. 106: 190–197.Google Scholar
  15. Gupta P.K., Roy J.K. and Prasad M. 2001. Single nucleotide polymorphisms: A new paradigm for molecular marker technology and DNA polymorphism detection with emphasis on their use in plants. Curr. Sci. 80: 524–535.Google Scholar
  16. Harry D.E., Temesgen B. and Neale D.B. 1998. Codominant PCR-based markers for Pinus taeda developed from mapped cDNA clones. Theor. Appl. Genet. 97: 327–336.Google Scholar
  17. Imyanitov E.N., Buslov K.G., Suspitsin E.N., Kuligina E., Belogubova E.V., Grigoriev M.Y., Togo A.V. and Hanson K.P. 2002. Improved reliability of allele-specific PCR. Biotechniques 33: 484–488.Google Scholar
  18. Jaramillo-Correa J.P., Beaulieu J. and Bousquet J. 2001. Contrasting evolutionary forces driving population structure at ESTPs, allozymes, and quantitative traits in white spruce. Mol. Ecol. 10:2729–2740.Google Scholar
  19. Kanazin V., Talbert H., See D., DeCamp P., Nevo E. and Blake T. 2002. Discovery and assay of single-nucleotide polymorphisms in barley Hordeum vulgare. Plant Mol. Biol. 48: 529–537.Google Scholar
  20. Kruglyak L. and Nickerson D.A. 2001. Variation is the spice of life. Nature Genet. 27: 234–236.Google Scholar
  21. Kwok P.Y., Carlson C., Yager T.D., Ankener W. and Nickerson D.A. 1994. Comparative analysis of human DNA variations by fluorescence-based sequencing of PCR products. Genomics 23: 138–144.Google Scholar
  22. Lai E., Riley J., Purvis I. and Roses A. 1998. A 4-Mb high-density single nucleotide polymorphism-based map around human APOE. Genomics 54: 31–38.Google Scholar
  23. Laroche J., Li P., Maggia L. and Bousquet J. 1997. Molecular evolution of angiosperm mitochondrial exons and introns. Proc. Nat. Acad. Sci. USA 94: 5722–5727.Google Scholar
  24. Latorra D., Hopkins D., Campbell K. and Hurley J.M. 2003. Multiple xallele-specific PCR with optimized locked nucleic acid primers. Biotechniques 34: 1150–1158.Google Scholar
  25. Miller K.M., Ming T.J., Schulze A.D. and Withler R.E. 1999. Denaturing gradient gel electrophoresis DGGE: A rapid and sensitive technique to screen nucleotide sequence variation in populations. Biotechniques 27: 1016–1030.Google Scholar
  26. Myers R.M., Maniatis T. and Lerman L.S. 1987. Detection and localization of single base changes by denaturing gradient gel electrophoresis. Meth. Enzymol. 155: 501–527.Google Scholar
  27. Nasu S., Suzuki J., Ohta R., Hasegawa K., Yui R., Kitazawa N., Monna L. and Minobe Y. 2002. Search for and analysis of single nucleotide polymorphisms SNPs in rice Oryza sativa, Oryzarufipogon and establishment of SNP markers. DNA Res. 9: 163–171.Google Scholar
  28. Neff M., Turk E. and Kalishman M. 2002. Web-based primer design for single nucleotide polymorphism analysis. Trends in Genetics 18: 613–615.Google Scholar
  29. Numakura C., Lin C., Ikegami T., Guldberg P. and Hayasaka K. 2002. Molecular analysis in Japanese patients with Charcot-Marie-Tooth disease: DGGE analysis for PMP22, MPZ, andCx32/GJB1 mutations. Hum. Mutat. 20: 392–398.Google Scholar
  30. Oefner P.J., Huber C.G., Umlauft F., Berti G.N., Stimpfl E. and Bonn G.K. 1994. High-resolution liquid chromatography of fluorescent dye-labeled nucleic acids. Anal. Biochem. 223: 39–46.Google Scholar
  31. Orita M., Iwahana H., Kanazawa H., Hayashi K. and Sekiya T. 1989. Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms. Proc. Natl. Acad. Sci. USA 86: 2766–2770.Google Scholar
  32. Paterson A.H., Bowers J.E., Burow M.D., Draye X., Elsik C.G., Jiang C.X., Katsar C.S., Lan T.H., Lin Y.R., Ming R.G. and Wright R.J. 2000. Comparative genomics of plant chromosomes. Plant Cell. 12: 1523–1539.Google Scholar
  33. Perron M., Perry D.J., Andalo C. and Bousquet J. 2000. Evidence from sequence-tagged-site markers of a recent progenitor-derivative species pair in conifers. Proc. Nat. Acad. Sci. USA 97: 11331–11336.Google Scholar
  34. Perry D.J. and Bousquet J. 1998a. Sequence-tagged-site STS markers of arbitrary genes: development, characterization and analysis of linkage in black spruce. Genetics 149: 1089–1098.Google Scholar
  35. Perry D.J. and Bousquet J. 1998b. Sequence-tagged-site STS markers of arbitrary genes: the utility of black spruce-derived STS primers in other conifers. Theor. Appl. Genet. 97: 735–743.Google Scholar
  36. Perry D.J. and Bousquet J. 2001. Genetic diversity and mating system of post-fire and post-harvest black spruce: an investigation using codominant sequence-tagged site STS markers. Can. J. For. Res. 31: 32–40.Google Scholar
  37. Perry D.J., Isabel N. and Bousquet J. 1999. Sequence-tagged-site STS markers of arbitrary genes: the amount and nature of variation revealed in Norway spruce. Heredity 83: 239–248.Google Scholar
  38. Picoult-Newberg L., Ideker T.E., Pohl M.G., Taylor S.L., Donald-son M.A., Nickerson D.A. and Boyce-Jacino M. 1999. MiningSNPs from EST databases. Genome Res. 9: 167–174.Google Scholar
  39. Plomion C., Hurme P., Frigerio J.M., Ridolfi M., Pot D., Pionneau C., Avila C., Gallardo F., David H., Neutelings G., Campbell M., Canovas F.M., Savolainen O., Bodenes C. and Kremer A. 1999. Developing SSCP markers in two Pinus species. Mol. Breed. 5: 21–31.Google Scholar
  40. Rickert A.M., Premstaller A., Gebhardt C. and Oefner P.J. 2002. Genotyping of SNPs in a polyploid genome by pyro sequencing. Biotechniques 32: 592–603.Google Scholar
  41. Sachidanandam R., Weissman D., Schmidt S.C., Kakol J.M., Stein L.D., Marth G., Sherry S., Mullikin J.C., Mortimore B.J., Willey D.L., Hunt S.E., Cole C.G., Coggill P.C., Rice C.M., Ning Z., Rogers J., Bentley D.R., Kwok P.Y., Mardis E.R., Yeh R.T., Schultz B., Cook L., Davenport R., Dante M., Fulton L., Hillier L., Waterston R.H., McPherson J.D., Gilman B., Schaffner S., Van Etten W.J., Reich D., Higgins J., Daly M.J., Blumenstiel B., Baldwin J., Stange-Thomann N., Zody M.C., Linton L., Lander E.S. and Altshuler D. 2001. A map of human genome sequence variation containing 1.42 million single nucleotide polymorphisms. Nature 409: 928–933.Google Scholar
  42. Schneider K., Weisshaar B., Borchardt D.C. and Salamini F. 2001. SNP frequency and allelic haplo type structure of Beta vulgari sexpressed genes. Mol. Breed. 8: 63–74.Google Scholar
  43. Schubert R., Mueller-Starck G. and Riegel R. 2001. Developmentof EST-PCR markers and monitoring their intra populational genetic variation in Picea abies L. Karst. Theor. Appl. Genet. 103: 1223–1231.Google Scholar
  44. Shaw S.H., Carrasquillo M.M., Kashuk C., Puffenberger E.G., and Chakravarti A. 1998. Allele frequency distributions in pooled DNA samples: applications to mapping complex disease genes. Genome Res. 8: 111–123.Google Scholar
  45. Shifman S., Pisante-Shalom A., Yakir B. and Darvasi A. 2002. Quantitative technologies for allele frequency estimation of SNPs in DNA pools. Mol. Cell. Probes 16: 429–434.Google Scholar
  46. Shubitowski D.M., Venta P.J., Douglass C.L., Zhou R.X. and Ewart S.L. 2001. Polymorphism identification within 50 equine gene-specific sequence tagged sites. Anim. Genet. 32: 78–88.Google Scholar
  47. Taillon-Miller P., Piernot E.E. and Kwok P.Y. 1999. Efficient approach to unique single-nucleotide polymorphism discovery. Genome Res. 9: 499–505.Google Scholar
  48. Temesgen B., Brown G.R., Harry D.E., Kinlaw C.S., Sewell M.M. and Neale D.B. 2001. Genetic mapping of expressed sequence tag polymorphism ESTP markers in loblolly pine Pinus taeda L. Theor. Appl. Genet. 102: 664–675.Google Scholar
  49. Tsumura Y., Suyama Y., Yoshimura K., Shirato N. and Mukai Y. 1997. Sequence-tagged-sites STSs of cDNA clones in Cryptomeriajaponica and their evaluation as molecular markers inconifers. Theor. Appl. Genet. 94: 764–772.Google Scholar
  50. Wang D.G., Fan J.B., Siao C.J., Berno A., Young P., Sapolsky R., Ghandour G., Perkins N., Winchester E., Spencer J., Kruglyak L., Stein L., Hsie L., Topaloglou T., Hubbell E., Robinson E., Mittmann M., Morris M.S., Shen N., Kilburn D., Rioux J., Nusbaum C., Rozen S., Hudson T.J., Lander E.S. 1998. Large-scale identification, mapping, and genotyping of single-nucleotide polymorphisms in the human genome. Science 280: 1077–1082.Google Scholar
  51. Wolford J.K., Blunt D., Ballecer C. and Prochazka M. 2000. High-throughput SNP detection by using DNA pooling and denaturing high performance liquid chromatography DHPLC. Hum. Genet. 107: 483–487.Google Scholar

Copyright information

© Kluwer Academic Publishers 2004

Authors and Affiliations

  1. 1.Chaire de recherche du Canada en génomique forestière et environnementale, Centre de recherche en biologie forestière, Pavillon Charles-Eugène-Marchand, Université LavalSainte-FoyCanada
  2. 2.Service Canadien des Forêts, Ressources naturelles Canada, Centre de foresterie des LaurentidesSainte-FoyCanada

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