Molecular Breeding

, Volume 21, Issue 3, pp 271–281

Application of multiplex-ready PCR for fluorescence-based SSR genotyping in barley and wheat

  • M. J. Hayden
  • T. M. Nguyen
  • A. Waterman
  • G. L. McMichael
  • K. J. Chalmers
Article

Abstract

Microsatellites (SSRs) are widely used in cereal research, and their use in marker assisted breeding has increased the speed and efficiency of germplasm improvement. Central to the application of SSRs for many purposes are methodologies enabling the low-cost acquisition of large quantities of genetic information for gene and genotype identification. In this study, multiplex-ready PCR was evaluated in barley and bread wheat as an approach for rapid and more automated SSR genotyping on a fluorescence-based DNA fragment analyzer. Multiplex-ready PCR is a method that allows SSR genotyping to be performed using a standardized protocol. The method enables flexible fluorescence labeling of SSRs, generates a relatively constant amount of PCR product for each marker, and has a high amenability to multiplex PCR (the simultaneous amplification of several SSRs in the same reaction). A high (92%) compatibility of published SSRs with multiplex-ready PCR is demonstrated, and the usefulness of the method for large scale genotyping is shown by its application for whole genome marker assisted breeding in barley. A database of more than 2,800 barley and wheat SSRs, and a suite of bio-informatic tools were developed to support the deployment of multiplex-ready PCR for various genetic applications, and are accessible at http://www.genica.net.au. Multiplex-ready PCR is broadly applicable to cereal genomics research and marker assisted breeding, and should be transferable to similar analyses of any animal or plant species.

Keywords

Marker assisted breeding Multiplex PCR Microsatellite SSR Semi-automated genotyping 

References

  1. Belgrader P, Marino MM, Lubin M, Baraby F (1996) A multiplex PCR-ligase detection reaction assay for human identity testing. Genome Sci Technol 1:77–87Google Scholar
  2. Bennett MD, Smith JB (1976) Nuclear DNA amounts in angiosperms. Philos Trans R Soc B 274:227–274CrossRefGoogle Scholar
  3. Blair MW, Hedetale V, McCouch SR (2002) Fluorescent-labeled microsatellite panels useful for detecting allelic diversity in cultivated rice (Oryza sativa L.). Theor Appl Genet 105:449–457PubMedCrossRefGoogle Scholar
  4. Crepieux S, Lebreton C, Flament P, Charmet G (2005) Application of a new IBD-based QTL mapping method to common wheat breeding population: analysis of kernel hardness and dough strength. Theor Appl Genet 111:1409–1419PubMedCrossRefGoogle Scholar
  5. Devos KM, Atkinson MD, Chinoy CN, Liu C, Gale MD (1992) RFLP-based genetic map of the homoeologous group-3 chromosomes of wheat and rye. Theor Appl Genet 83:931–939CrossRefGoogle Scholar
  6. Donini P, Stephenson P, Bryan GJ, Koebner RMD (1998) The potential of microsatellites for high throughput genetic diversity assessment in wheat and barley. Genet Resour Crop Evol 45:415–421CrossRefGoogle Scholar
  7. Edwards MC, Gibbs RA (1994) Multiplex PCR: advantages, development and applications. PCR Methods Appl 3:S65–S75PubMedGoogle Scholar
  8. Gupta PK, Rustgi S (2004) Molecular markers from the transcribed/expressed region of the genome in higher plants. Funct Integr Genomics 4:139–162PubMedCrossRefGoogle Scholar
  9. Gupta PK, Varshney RK (2000) The development of use of microsatellite markers for genetic analysis and plant breeding with emphasis on bread wheat. Euphytica 113:163–185CrossRefGoogle Scholar
  10. Gupta PK, Varshney RK, Sharma PC, Ramesh B (1999) Molecular markers and their application in wheat breeding. Plant Breed 118:369–390CrossRefGoogle Scholar
  11. Hearnden PR, Eckermann PJ, McMichael GL, Hayden MJ, Eglinton JK, Chalmers KJ (2007) A genetic map of 1000 SSR and DArT loci in a wide barley cross. Theor Appl Genet. doi:10.1007/s00122-007-0572-7Google Scholar
  12. Heller C (2001) Principles of DNA separation with capillary electrophoresis. Electrophoresis 22:629–643PubMedCrossRefGoogle Scholar
  13. Henegariu O, Heerema NA, Dlouhy SR, Vance GH, Vogt PH (1997) Multiplex PCR: critical parameters and step-by-step protocol. Biotechniques 23:504–511PubMedGoogle Scholar
  14. Koebner RMD, Summers RW (2003) 21st century wheat breeding: plot selection or plate selection? Trends Biotech 21:59–63CrossRefGoogle Scholar
  15. Langridge P, Barr AR (2003) Better barley faster: the role of marker assisted selection—preface. Aust J Agric Res 54:1–5CrossRefGoogle Scholar
  16. Macaulay M, Ramsay L, Powel W, Waugh R (2001) A representative, highly informative ‘genotyping set’ of barley SSRs. Theor Appl Genet 102:801–809CrossRefGoogle Scholar
  17. Mansfield DC, Brown AF, Green DK, Carothers AD, Morris SW, Evans H-J (1994) Automation of genetic linkage analysis using fluorescent microsatellite markers. Genomics 24:225–233PubMedCrossRefGoogle Scholar
  18. Masi O, Spagnoletti-Zeuli PL, Donini P (2003) Development and analysis of multiplex microsatellite marker sets in common bean (Phaseolus vulgaris L.). Mol Breed 11:303–313CrossRefGoogle Scholar
  19. McCouch SR, Teytelman L, Xu Y, Lobos KB, Clare K, Walton M, Fu B, Maghirang R, Li Z, Xing Y, Zhang Q, Kono I, Yano M, Fjellstrom R, DeClerck G, Schneider D, Cartinhour S, Ware D, Stein L (2002) Development and mapping of 2240 new SSR markers for rice (Oryza sativa L.). DNA Res 9:199–207PubMedCrossRefGoogle Scholar
  20. Mitchell SE, Kresovich S, Jester CA, Hernandez CJ, Szewc-McFadden AK (1997) Application of multiplex PCR and fluorescence-based, semi-automated allele sizing technology for genotyping plant genetic resources. Crop Sci 37:617–624CrossRefGoogle Scholar
  21. Narvel JM, Chu WC, Fehr WR, Cregan PB, Shoemaker RC (2000) Development of multiplex sets of simple sequence repeat DNA markers covering the soybean genome. Mol Breed 6:175–183CrossRefGoogle Scholar
  22. Oetting WS, Lee HK, Flanders DJ, Wiesner GL, Sellers TA, King RA (1995) Linkage analysis with multiplexed short tandem repeat polymorphisms using infared fluorescence and M13 tailed primers. Genomics 30:450–458PubMedCrossRefGoogle Scholar
  23. Paris M, Carter M (2000) Cereal DNA: a rapid high-throughput extraction method for marker assisted selection. Plant Mol Biol Rep 18:357–360CrossRefGoogle Scholar
  24. Perry DJ (2004) Identification of Canadian durum wheat varieties using a single PCR. Theor Appl Genet 109:55–61PubMedCrossRefGoogle Scholar
  25. Ponce MR, Robles P, Micol JL (1999) High-throughput genetic mapping in Arabidopsis thalina. Mol Gen Genet 261:408–415PubMedCrossRefGoogle Scholar
  26. Prasad M, Varshney RK, Roy JK, Balyan HS (2000) The use of microsatellites for detecting DNA polymorphisms, genotype identification and genetic diversity in wheat. Theor Appl Genet 100:584–592Google Scholar
  27. Ramsay L, Macaulay M, Ivanissevich DS, MacLean K, Cardle L, Fuller J, Edwards KJ, Tuvesson S, Morgante M, Masssari A, Maestri E, Marmiroli N, Sjakste T, Ganal M, Powell W, Waugh R (2000) A simple sequence repeat-based linkage map of barley. Genetics 156:1997–2005PubMedGoogle Scholar
  28. Roder MS, Korzun V, Wendehake K, Plaschke J, Tixier M-H, Leroy P, Ganal MW (1998) A microsatellite map of wheat. Genetics 149:2007–2023PubMedGoogle Scholar
  29. Sambrook J, Russell (2001) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
  30. Sharopova N, McMullen MS, Schultz L, Schroeder S, Sanchez-Villeda H, Gardiner J, Bergstrom D, Houchins K, Melia-Hancock S, Musket T, Duru N Polacco M, Edwards K, Ruff T, Register JC, Brouwer C, Thompson R, Velasco R, Chin E, Lee M, Woodman-Clikeman W, Long MJ, Liscum E, Cone K, Davis G, Coe EH Jr (2002) Development and mapping of SSRs in maize. Plant Mol Biol 48:463–481PubMedCrossRefGoogle Scholar
  31. Somers DJ, Isaac P, Edwards K (2004) A high-density microsatellite consensus map for bread wheat (Triticum aestivum L.). Theor Appl Genet 109:1105–1114PubMedCrossRefGoogle Scholar
  32. Song QJ, Shi JR, Singh S, Fickus EW, Costa JM, Lewis J, Gill GS, Ward R, Cregan PB (2005) Development and mapping of microsatellite (SSR) markers in wheat. Theor Appl Genet 110:550–560PubMedCrossRefGoogle Scholar
  33. Stein N, Graner A (2004) Map-based gene isolation in cereal genomes. In: Gupta PK, Varshney RK (eds) Cereal genomics. Kluwer, Dordrecht, pp 331–360Google Scholar
  34. Tang S, Kishore VK, Knapp SJ (2003) PCR-multiplexes for a genome-wide framework of simple sequence repeat marker loci in cultivated sunflower. Theor Appl Genet 107:6–19PubMedGoogle Scholar
  35. Tommasini L, Batley J, Arnold GM, Cooke RJ, Donini P, Lee D, Law JR, Lowe C, Moule C, Trick M, Edwards KJ (2003) The development of multiplex simple sequence repeats (SSR) markers to complement distinctness, uniformity and stability testing of rape (Brassica napus L.) varieties. Theor Appl Genet 106:1091–1101PubMedGoogle Scholar
  36. Varshney RK, Sigmund R, Borner A, Korzun V, Stein N, Sorrells ME, Langridge P, Graner A (2005) Interspecific transferability and comparative mapping of barley EST-SSR markers in wheat, rye and rice. Plant Sci 168:195–202CrossRefGoogle Scholar
  37. Varshney RK, Marcel TA, Ramsay L, Russel J, Roder MS, Stein N, Waugh R, Langridge P, Niks RE, Graner A (2007) A high density barley microsatellite consensus map with 775 SSR loci. Theor Appl Genet 114:1091–1103PubMedCrossRefGoogle Scholar
  38. Vos P, Hogers R, Bleeker M, Reijans M, van de Lee T, Hornes M, Frijters A, Pot J, Peleman J, Kuiper M, Zabeau M (1995) AFLP: a new technique for DNA fingerprinting. Nucleic Acids Res 23:4007–4414CrossRefGoogle Scholar
  39. Ziegle JS, Su Y, Corcoran KP, Nie L, Maynard PE, Hoff LB, McBride LJ, Kronick MN, Diehl SR (1992) Application of automated DNA sizing technology for genotyping microsatellite loci. Genomics 14:1026–1031PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2007

Authors and Affiliations

  • M. J. Hayden
    • 1
    • 2
  • T. M. Nguyen
    • 1
    • 2
  • A. Waterman
    • 1
    • 2
  • G. L. McMichael
    • 1
    • 2
  • K. J. Chalmers
    • 1
    • 2
  1. 1.Molecular Plant Breeding CRCGlen OsmondAustralia
  2. 2.School of Agriculture, Food and WineUrrbraeAustralia

Personalised recommendations