Functional & Integrative Genomics

, Volume 8, Issue 3, pp 223–233

EST-derived single nucleotide polymorphism markers for assembling genetic and physical maps of the barley genome

  • R. Kota
  • R. K. Varshney
  • M. Prasad
  • H. Zhang
  • N. Stein
  • A. Graner
Original Paper


In a panel of seven genotypes, 437 expressed sequence tag (EST)-derived DNA fragments were sequenced. Single nucleotide polymorphisms (SNPs) that were polymorphic between the parents of three mapping populations were mapped by heteroduplex analysis and a genome-wide consensus map comprising 216 EST-derived SNPs and 4 InDel (insertion/deletion) markers was constructed. The average frequency of SNPs amounted to 1/130 bp and 1/107.8 bp for a set of randomly selected and a set of mapped ESTs, respectively. The calculated nucleotide diversities (π) ranged from 0 to 40.0 × 10−3 (average 3.1 × 10−3) and 0.52 × 10−3 to 39.51 × 10–3 (average 4.37 × 10−3) for random and mapped ESTs, respectively. The polymorphism information content value for mapped SNPs ranged from 0.24 to 0.50 with an average of 0.34. As expected, combination of SNPs present in an amplicon (haplotype) exhibited a higher information content ranging from 0.24 to 0.85 with an average of 0.50. Cleaved amplified polymorphic sequence assays (including InDels) were designed for a total of 87 (39.5%) SNP markers. The high abundance of SNPs in the barley genome provides avenues for the systematic development of saturated genetic maps and their integration with physical maps.


Molecular markers SNPs Haplotype diversity Nucleotide diversity Genetic map 

Supplementary material

Table ESM 1Details on development and characterization of SNP markers (HTML 360 kb)
10142_2007_60_MOESM2_ESM.doc (83 kb)
Table ESM 2Informative set of SNP markers for haplotype analysis and CAPS assays (DOC 83 kb)


  1. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410PubMedGoogle Scholar
  2. Antonarakis SE (1998) Recommendations for a nomenclature system for human gene mutations. Human Mutat 11:1–3CrossRefGoogle Scholar
  3. Batley J, Barker G, O'Sullivan H, Edwards KJ, Edwards D (2003) Mining for single nucleotide polymorphisms and insertions/deletions in maize expressed sequence tag data. Plant Physiol 132:84–91PubMedCrossRefGoogle Scholar
  4. Beutler E, McKusick VA, Motulsky AG, Scriver CR, Hutchinson F (1996) Mutation nomenclature: nicknames, systematic names, and unique identifiers. Human Mutat 8:203–206CrossRefGoogle Scholar
  5. Bundock PC, Christopher JT, Eggler P, Ablett G, Henry RJ, Holton TA (2003) Single nucleotide polymorphisms in cytochrome P450 genes from barley. Theor Appl Genet 106:676–682PubMedGoogle Scholar
  6. Bundock PC, Henry RJ (2004) Single nucleotide polymorphism, haplotype diversity and recombination in the Isa gene of barley. Theor Appl Genet 109:543–551PubMedCrossRefGoogle Scholar
  7. Chiapparino E, Lee D, Donini P (2004) Genotyping single nucleotide polymorphisms in barley by tetra-primer ARMS-PCR. Genome 47:414–420PubMedGoogle Scholar
  8. Ching A, Caldwell KS, Jung M, Dolan M, Smith OS, Tingey S, Morgante M, Rafalski AJ (2002) SNPs frequency, halpotype structure and linkage disequilibrium in elite maize inbred lines. BMC Genetics 3:19PubMedCrossRefGoogle Scholar
  9. Cho RJ, Mindrinos M, Richards DR, Sapolsky RJ, Sapolsky RJ, Anderson M, Drenkard E, Dewdney L, Reuber TL, Stammers M, Federspiel N, Theologis A, Yang WH, Hubbell E, Au M, Chung EY, Lashkari D, Lemieux B, Dean C, Lipshutz RJ, Ausubel FM, Davis RW, Oefner PJ (1999) Genome-wide mapping with biallelic markers in Arabidopsis thaliana. Nature Genet 23:203–207PubMedCrossRefGoogle Scholar
  10. Costa JM, Corey A, Hayes PM, Jobet C, Kleihofs A et al (2001) Molecular mapping of the Oregon Wolfe Barleys: a phenotypically polymorphic doubled-haploid population. Theor Appl Genet 103:415–424CrossRefGoogle Scholar
  11. den Dunnen JT, Antonarakis SE (2000) Mutation nomenclature extensions and suggestions to describe complex mutations: A discussion. Theor Appl Genet 15:7–12Google Scholar
  12. Ewing B, Hillier L, Wendl MC, Green P (1998) Base-calling of automated sequencer traces using phred. I. Accuracy assessment. Genome Res 8:175–185Google Scholar
  13. Feltus FA, Wan J, Schulze SR, Estill JC, Jiang N, Paterson AH (2004) An SNPs resource for rice genetics and breeding based on subspecies indica and japonica genome alignments. Genome Res 14:1812–1819PubMedCrossRefGoogle Scholar
  14. Graner A, Jahoor A, Schondelmaier H, Siedler K, Pillen K, Wenzel G, Herrmann RG (1991) Construction of an RFLP map of barley. Theor Appl Genet 83:250–256CrossRefGoogle Scholar
  15. Graner A, Dehmer KJ, Thiel T, Börner A (2004) Plant genetic resources: benefits and implications of using molecular markers. In: Carmen de Vincente M (ed) Issues in genetic resources no. 11. IPGRI, Rome, pp 26–32Google Scholar
  16. Gribskov M, Devereux J, Burgess RR (1984) The codon preference plot: graphic analysis of protein coding sequences and prediction of gene expression. Nucleic Acids Res 12:539–549PubMedCrossRefGoogle Scholar
  17. Hartl DL, Clark AG (1997) Principles of population genetics. Sinauer Associates, Sunderland, USAGoogle Scholar
  18. Hamblin MT, Mitchell SE, White GM, Gallego J, Kukatla R, Wing RA, Paterson AH, Kresovich S (2004) Comparative population genetics of the panicoid grasses: sequence polymorphism, linkage disequilibrium and selection in a diverse sample of Sorghum bicolor. Genetics 167:471–483PubMedCrossRefGoogle Scholar
  19. Kanazin V, Talbert H, See D, Decamp P, Nevo E, Blake T (2002) Discovery and assay of single nucleotide polymorphism in barley (Hordeum vulgare). Plant Mol Biol 48:529–537PubMedCrossRefGoogle Scholar
  20. Kleinhofs A, Graner A (2001) An integrated map of the barley genome. In: Phillips RL, Vasil IK (ed) DNA markers in plants. Kluwer, Dordrecht, The Netherlands, pp 187–199Google Scholar
  21. Kleinhofs A, Kilian A, Saghai Maroof M, Biyashev R, Hayes P, Chen FQ, Lapitan N, Fenwick A, Blake TK, Kanazin V, Ananiev E, Dahleen L, Kudrna D, Bollinger J, Knapp SJ, Liu B, Sorrells M, Heun M, Franckowiak JD, Hoffman D, Skadsen R, Steffenson BJ (1993) A molecular isozyme and morphological map of barley (Hordeum vulgare) genome. Theor Appl Genet 86:705–712CrossRefGoogle Scholar
  22. Kota R, Varshney RK, Thiel T, Dehmer K-J, Graner A (2001a) Generation and comparison of EST-derived SSR and SNPs markers in barley (Hordeum vulgare L.). Hereditas 135:141–151CrossRefGoogle Scholar
  23. Kota R, Wolf M, Michalek W, Graner A (2001b) Application of DHPLC for mapping of single nucleotide polymorphisms (SNPs) in barley (Hordeum vulgare L.). Genome 44:523–528PubMedCrossRefGoogle Scholar
  24. Kota R, Rudd S, Facius A, Kolesov G, Thiel T, Zhang H, Stein N, Mayer K, Graner A (2003) Snipping polymorphisms from large EST collections in barley (Hordeum vulgare L.). Mol Genet Genomics 270:24–33PubMedCrossRefGoogle Scholar
  25. Möhring S, Salamini F, Schneider K (2004) Multiplexed, linkage group-specific marker sets for rapid genetic mapping and fingerprinting of sugar beet (Beta vulgaris L.). Mol Breed 14:475–488CrossRefGoogle Scholar
  26. Nasu S, Suzuki J, Ohta R, Hasegawa K, Yui R, Kitazawa N, Monna L, Minobe Y (2002) Search for and analysis of single nucleotide polymorphisms (SNPs) in rice (Oryza sativa, Oryza rufipogon) and establishment of SNPs markers. DNA Res 9:163–171PubMedCrossRefGoogle Scholar
  27. Nei M (1987) Molecular Evolutionary Genetics. Columbia University Press, NewYork USAGoogle Scholar
  28. Nei M, Li WH (1979) Mathematical model for studying genetic variation in terms of restriction endonucleases. Proc Natl Acad Sci U S A 76:5269–5273PubMedCrossRefGoogle Scholar
  29. Picoult-Newberg L, Ideker TE, Pohl MG, Taylor SL, Donaldson MA, Nickerson DA, Boyce-Jacino M (1999) Mining SNPs from EST databases. Genome Res 9:167–174PubMedGoogle Scholar
  30. Qi LL, Echalier B, Chao S, Lazo GR, Butler GE, Anderson OD, Akhunov ED, Dvorak J, Linkiewicz AM et al (2004) A chromosome bin map of 16,000 expressed sequence tag loci and distribution of genes among the three genomes of polyploid wheat. Genetics 168:701–712PubMedCrossRefGoogle Scholar
  31. Rafalski JA (2002) Application of single nucleotide polymorphisms in crop genetics. Curr Opin Plant Biol 5:94–100PubMedCrossRefGoogle Scholar
  32. Rostoks N, Mudie S, Cardle L, Russell J, Ramsay L, Booth A, Svensson JT, Wanamaker SI, Walia H, Rodriguez EM, Hedley PE, Liu H, Morris J, Close TJ, Marshall DF, Waugh R (2005) Genome-wide SNPs discovery and linkage analysis in barley based on genes responsive to abiotic stress. Mol Genet Genomics 274:515–527PubMedCrossRefGoogle Scholar
  33. Rostoks N, Ramsay L, MacKenzie K, Cardle L, Svensson JT, Prasanna B, Stein N, Varshney RK, Marshall D, Graner A, Close TJ, Waugh R (2006) Recent history of artificial outcrossing facilitates whole-genome association mapping in elite inbred crop varieties. Proc Natl Acad Sci 103:18656–18661PubMedCrossRefGoogle Scholar
  34. Russell J, Fuller J, Macaulay M, Hatz BG, Jahoor A, Powell W, Waugh R (1997) Direct comparison of levels of genetic variation among barley accessions detected by RFLPs, AFLPs, SSRs and RAPDs. Theor Appl Genet 9:714–722CrossRefGoogle Scholar
  35. Russell J, Booth A, Fuller J, Harrower B, Hedley P, Machray G, Powell W (2004) A comparison of sequence-based polymorphism and haplotype content in transcribed and anonymous regions of the barley genome. Genome 47:389–398PubMedCrossRefGoogle Scholar
  36. Sachidanandam R, Weissman D, Schmidt SC, Kakol JM, Stein LD, Mullikin JC, Mortimore BJ, Willey DL, Hunt SE, Cole CG (2001) A map of human genome sequence variation containing 1.42 million single nucleotide polymorphisms. Nature 409:928–933PubMedCrossRefGoogle Scholar
  37. Schmid KJ, Sorensen TR, Stracke R, Torjek O, Altmann T, Mitchell-Olds T, Weisshaar B (2003) Large-scale identification and analysis of genome-wide single-nucleotide polymorphisms for mapping in Arabidopsis thaliana. Genome Res 13:1250–1257PubMedCrossRefGoogle Scholar
  38. Schneider K, Weisshaar B, Borchardt DC, Salamini F (2001) SNPs frequency and allelic haplotype of Beta vulgaris expressed genes. Mol Breed 8:63–74CrossRefGoogle Scholar
  39. Somers DJ, Kirkpatrick R, Moniwa M, Walsh A (2003) Mining single-nucleotide polymorphisms from hexaploid wheat ESTs. Genome 46:431–437PubMedCrossRefGoogle Scholar
  40. Stam P (1993) Construction of integrated genetic linkage maps by means of a new computer package: JoinMap. Plant J 3:739–744CrossRefGoogle Scholar
  41. Stein N, Prasad M, Scholz U, Thiel T, Zhang H, Wolf M, Kota R, Varshney RK, Perovic D, Grosse I, Graner A (2007) A 1000 loci transcript map of the barley genome—new anchoring points for integrative grass genomics. Theor Appl Genet 114:823–839PubMedCrossRefGoogle Scholar
  42. Tenaillon MI, Sawkins MC, Long AD, Gaut B, Doebley JF, Brandon S (2001) Patterns of DNA sequence polymorphism along chromosome 1 of maize (Zea mays ssp. Mays L.). Proc Natl Acad Sci U S A 98:9161–9166PubMedCrossRefGoogle Scholar
  43. Thiel T, Michalek W, Varshney RK, Graner A (2003) Exploiting EST databases for the development of cDNA derived microsatellite markers in barley (Hordeum vulgare L.). Theor Appl Genet 106:411–422PubMedGoogle Scholar
  44. Thiel T, Kota R, Grosse I, Stein N, Graner A (2004) SNP2CAPS: a SNPs and INDEL analysis tool for CAPS marker development. Nucleic Acids Res 32(1):e5PubMedCrossRefGoogle Scholar
  45. Thompson JD, Higgins DG, Gibson TJ (1994) Clustal-W—improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680PubMedCrossRefGoogle Scholar
  46. Torjek O, Berger D, Meyer RC, Mussig C, Schmid KJ, Rosleff Sorensen T, Weisshaar B, Mitchell-Olds T, Altmann T (2003) Establishment of a high-efficiency SNPs-based framework marker set for Arabidopsis. Plant J 36:122–140PubMedCrossRefGoogle Scholar
  47. Van K, Hwang E-Y, Young Kim M, Kim Y-H, Cho Y-I, Cregan PB, Lee S-H (2004) Discovery of single nucleotide polymorphisms in soybean using primers designed from ESTs. Euphytica 139:147–157CrossRefGoogle Scholar
  48. Varshney RK, Prasad M, Graner A (2004) Molecular marker maps of barley: a resource for intra- and interspecific genomics. In: Wenzel G, Horst L (eds) Molecular markers in improvement of agriculture and forestry, Springer, Germany, pp 229–243Google Scholar
  49. Varshney RK, Grosse I, Hahnel U, Siefken R, Prasad M, Stein N, Langridge P, Altschmied L, Graner A (2006) Genetic mapping and BAC assignment of EST-derived SSR markers shows non-uniform distribution of genes in the barley genome. Theor Appl Genet 113:239–250PubMedCrossRefGoogle Scholar
  50. Varshney RK, Beier U, Khlestkina E, Kota R, Korzun V, Röder M, Graner A, Börner A (2007) Single nucleotide polymorphisms in rye: discovery, frequency and applications for genome mapping and diversity studies. Theor Appl Genet 114:1105–1116PubMedCrossRefGoogle Scholar
  51. Zhang H, Sreenivasulu N, Weschke W, Stein N, Rudd S, Radchuk V, Potokina E, Scholz U, Schweizer P, Zierold U, Langridge P, Varshney RK, Wobus U, Graner A (2004) Large-scale analysis of the barley transcriptome based on expressed sequence tags. Plant J 40:276–290PubMedCrossRefGoogle Scholar
  52. Zhu YL, Song QJ, Hyten DL, van Tassell C, Matukumalli LK, Grimm DR, Hyatt SM, Fickus EW, Young ND, Cregan PB (2003) Single-nucleotide polymorphisms in soybean. Genetics 163:1123–1134PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • R. Kota
    • 1
    • 5
  • R. K. Varshney
    • 2
    • 5
  • M. Prasad
    • 3
    • 5
  • H. Zhang
    • 4
    • 5
  • N. Stein
    • 5
  • A. Graner
    • 5
  1. 1.Plant Disease Resistance GroupCSIRO–Plant IndustryCanberraAustralia
  2. 2.International Crops Research Institute for the Semi-Arid Tropics (ICRISAT)PatancheruIndia
  3. 3.National Institute for Plant Genome Research (NIPGR)New DelhiIndia
  4. 4.Laboratory of Molecular Plant PhysiologyUniversity of FloridaGainesvilleUSA
  5. 5.Leibniz Institute of Plant Genetics and Crop Plant Research (IPK)GaterslebenGermany

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