Theoretical and Applied Genetics

, Volume 107, Issue 6, pp 1113–1122 | Cite as

Grass consensus STS markers: an efficient approach for detecting polymorphism in Lolium



For ryegrass and many forage crops, characterization of varieties is often difficult for two reasons: few of discriminant morphological traits and a great within-varieties variation. Futhermore, few molecular markers are publicly accessible. In this paper we describe two approaches for the development of 42 sequence-tagged-site (STS) markers. Firstly, 14 STS markers were developed from Lolium sequences found in data bases. Secondly, 28 STS markers were developed from sequences found in related species of Gramineae. Out of 42 STS markers developed, 85.8% yielded successfull amplification and 62% revealed a high level of polymorphism with an average of five alleles per locus. The analysis of amplicons reveals a high STS marker specificity, a high conservation in gene structure and a strong intron sequence homology between allelic forms. Moreover, the majority of the STS markers can be considered as "universal markers" because 81% of these STS markers amplified successfully across 20 related grass species. These results permit us to consider the use of these markers in synteny studies.


Lolium STS Consensus marker Grass species Intronic polymorphism 



This project was partly supported by financial assistance from CTPS (Comité Technique Permanent de la Sélection) and CIRAD (Centre de Coopération Internationale en Recherche Agronomique pour le développement. France). We also thank the CIRAD for providing the rice and sugarcane DNAs. We are very grateful to Virginie Lauvergeat for her STS sequencing results and her valuable comments on this manuscript.


  1. Balfourier F, Charmet G (1994) Geographical patterns of isozyme variation in Mediterranean populations of perennial ryegrass. Heredity 72:55–63Google Scholar
  2. Bert PF, Charmet G, Sourdille P, Hayward MD, Balfourier F (1999) A high-density molecular map for ryegrass (Lolium perenne) using AFLP markers. Theor Appl Genet 99:445–452CrossRefGoogle Scholar
  3. Chalhoub BA, Thibault S, Laucou V, Rameau C, Hofte H, Cousin R (1997) Silver staining and recovery of AFLP amplification products on large denaturing polyacrylamide gels. Biotechniques 22:216–220PubMedGoogle Scholar
  4. Chen HB, Martin JM, Lanvin M, Talbert LE (1994) Genetic diversity in hart red spring wheat based on sequence-tagged-site PCR markers. Crop Sci 34:1628–1632Google Scholar
  5. Cheung WY, Hubert N, Landry BS (1993) A simple and rapid DNA microextraction method for plant, animal and insects suitable for RAPD and other PCR analyses. PCR Methods Appl 3:69–70PubMedGoogle Scholar
  6. Clark AG, Muse SV, Leicht BG (1996) Length variation and secondary structure of introns in the Mlc1 gene in six species of Drosophila. Mol Biol Evol 13:471–482PubMedGoogle Scholar
  7. Cosgrove DJ, Bedinger P, Durachko DM (1997) Group I allergens of grass pollen as cell wall-loosening agents. Proc Natl Acad Sci USA 94:6559–6564CrossRefPubMedGoogle Scholar
  8. Frugoli JA, McPeek MA, Thomas TL, McClung CR (1998) Intron loss and gain during evolution of the catalase gene family in angiosperms. Genetics 149:355–365PubMedGoogle Scholar
  9. Gaut BS, Peek AS, Morton BR, Clegg MT (1999) Patterns of genetic diversification within the Adh gene family in the grasses (Poaceae). Mol Biol Evol 16:1086–1097PubMedGoogle Scholar
  10. Ghareyazie B, Huang N, Second G, Bennett J, Khush GS (1995) Classification of rice germplasm. I. Analysis using ALP and PCR-based RFLP. Theor Appl Genet 91:218–227Google Scholar
  11. Gilpin BJ, McCallum JA, Frew TJ, Timmerman-Vaughan GM (1997) A linkage map of the pea (Pisum sativum L.) genome containing cloned sequences of known function and expressed sequence tags (ESTs). Theor Appl Genet 95:1289–1299CrossRefGoogle Scholar
  12. Greneche M, Lallemand J, Michaud O (1991) Comparaison of different enzyme loci as a means of distinguishing ryegrass varieties by electrophoresis. Seed Sci Technol 19:147–158Google Scholar
  13. Hayward MD, McAdam NJ (1977) Isozyme polymorphism as a measure of distinctness and stability in cultivars of L. perenne. Z PflanzenZücht 79:59–68Google Scholar
  14. Jones ES, Dupal MP, Kölliker R, Drayton MC, Forster JW (2001) Development and characterisation of simple sequence repeat (SSR) markers for perennial ryegrass (Lolium perenne L.). Theor Appl Genet 102:405–415Google Scholar
  15. Jones ES, Mahoney NL, Hayward MD, Armstead IP, Jones JG, Humphreys MO, King IP, Kishida T, Yamada T, Balfourier F, Charmet G, Forster JW (2002) An enhanced molecular marker-based genetic map of perennial ryegrass (Lolium perenne) reveals comparative relationships with other Poaceae genomes. Genome 45:282–295PubMedGoogle Scholar
  16. Kirby DA, Muse SV, Stephan W (1995) Maintenance of pre-mRNA secondary structure by epistatic selection. Proc Natl Acad Sci USA 92:9047–9051PubMedGoogle Scholar
  17. Kubik C, Meyer WA, Gaut BS (1999) Assessing the abundance and polymorphism of simple sequence repeats in perennial Ryegrass. Crop Sci 39:1136–1141Google Scholar
  18. Kubik C, Sawkins M, Meyer WA, Gaut BS (2001) Genetic diversity in seven perennial ryegrass (Lolium perenne L.) cultivars based on SSR markers. Crop Sci 41:1565–1572Google Scholar
  19. Lallemand J, Lem P, Ghesquière M, Charmet G, Balfourier F (1998) Potentiality of STSs for variety distinction in ryegrass. In: UPOV (ed) Working group on biochemical and molecular techniques and DNA-profiling in particular. Session 5, Betsville, USA, September 28–30, 1998, BMT/S11Google Scholar
  20. Leicht BG, Muse SV, Hanczyc M, Clark AG (1995) Constraints on intron evolution in the gene encoding the myosin alkali light chain in Drosophila. Genetics 139:299–308PubMedGoogle Scholar
  21. Mitchell LE, Dennis ES, Peacock WJ (1989) Molecular analysis of an alcohol dehydrogenase (Adh) gene from chromosome 1 of wheat. Genome 32:349–358PubMedGoogle Scholar
  22. Rogers SO, Bendich AJ (1988) Extraction of DNA from plant tissues. Plant Mol Biol A6:1–10Google Scholar
  23. Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA (1988) Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239:487–491PubMedGoogle Scholar
  24. Talbert LE, Blake NK, Chee PW, Blake TK, Magyar GM (1994) Evaluation of "sequence-tagged-site" PCR products as molecular markers in wheat. Theor Appl Genet 87:789–794Google Scholar
  25. Taylor C, Madsen K, Borg S, Moller MG, Boelt B, Holm PB (2001) The development of sequence-tagged sites (STSs) in Lolium perenne L.: the application of primer sets derived from other genera. Theor Appl Genet 103:648–658Google Scholar
  26. Tragoonrung S, Kanazin V, Hayes PM, Blake TK (1992) Sequence-tagged-site-facilitated PCR for barley genome mapping. Theor Appl Genet 84:1002–1008Google Scholar

Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  1. 1.BioGEVES, INRA du Magneraud, BP 52, 17700 Surgères, France

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