, Volume 209, Issue 2, pp 507–523 | Cite as

Models with only two predictor variables can accurately predict seed yield in diploid and tetraploid red clover

  • Tim Vleugels
  • Bob Ceuppens
  • Gerda Cnops
  • Peter Lootens
  • Frederik R. D. van Parijs
  • Guy Smagghe
  • Isabel Roldán-Ruiz


Red clover (Trifolium pratense L.) is a valuable protein-rich forage crop, but poor seed yields are often a setback, especially in tetraploid varieties. Published work on factors underlying seed yield report contradictory or (sometimes) inconclusive results. Here we report on the effect of 10 traits related to seed yield and flowering on seed yield in 600 genotypes from 15 diploid and 15 tetraploid cultivars from diverse origins. Multiple linear regression (MLR) models indicated that the variation in seed number per plant was well explained by (1) the number of ripe flower heads per plant and (2) the seed number per ripe flower head in diploids (R2 = 0.953) and tetraploids (R2 = 0.919). Flower color was not significantly related to seed yield, and corolla tube dimensions were not (or only borderline) significantly related to seed yield. The applicability of our models to predict seed yield was validated on the same set of plants in the following harvest year, and on a totally different dataset available from Vleugels et al. (Plant Breed 134:56–61, 2015). In the 600-plant dataset and the separate Vleugels et al. dataset, the diploid model explained 94.8 and 53.2 %, respectively, and the tetraploid model explained 88.1 and 64.9 %, respectively, of the variation in seed yield. Our models can therefore predict seed yield with good to fair accuracy in red clover plants grown under various conditions. Breeders can increase seed yield by selecting plants that have numerous ripe flower heads and a high seed number per head. Flower color and corolla tube dimensions can be excluded as breeding targets.


Trifolium pratense L. Seed yield Flower morphology Corolla tube dimensions 



Anthocyanin content


Corolla tube diameter


Corolla tube length




Flower number/flower head


Total flower length


Principal component analysis


Multiple linear regression


Number of ripe flower heads/plant


Seed number/plant


Seed weight/plant




Number of unripe flower heads/plant



We would like to thank Nancy Mergan, Katleen Sucaet, Thomas Vanderstocken, Luc Hertegonne, Greta Simoens, Ariane Staelens, Carina Pardon, Geert Lejeune, Luc Van Ghysegem, Kurt Snaet, Filip Coussens and Johan Snoeck for their assistance in measuring the flower morphology, harvesting seeds, and field trial maintenance. We thank Dr. ir. Marie-Christine van Labeke (Ghent University, Belgium) for her recommendations on spectrophotometry, and Dr. Beat Boller (Agroscope, Switzerland) for his advice on our set of cultivars.

Supplementary material

10681_2016_1679_MOESM1_ESM.docx (43 kb)
Supplementary material 1 (DOCX 43 kb)
10681_2016_1679_MOESM2_ESM.docx (26 kb)
Supplementary material 2 (DOCX 25 kb)


  1. Adamse P, Peters JL, Jaspers P, van Tuinen A, Koornneef M, Kendrick RE (1989) Photocontrol of anthocyanin synthesis in tomato seedlings: a genetic approach. Photochem Photobiol 50:107–111CrossRefGoogle Scholar
  2. Amdahl H, Aamlid ST, Ergon A, Kovi MR, Marum P, Alseikh M, Rognli OA (2015) Seed yield of Norwegian and Swedish tetraploid red clover (Trifolium pratense L.) populations. Crop Sci (in press). doi: 10.2135/cropsci2015.07.0441
  3. Annicchiarico P, Barrett B, Brummer EC, Julier B, Marshall AH (2014) Achievements and challenges in improving temperate perennial forage legumes. Crit Rev Plant Sci 34:327–380CrossRefGoogle Scholar
  4. Barrett BA, Baird IJ, Woodfield DR (2004) Genetic tools for increased white clover seed production. Proc New Zeal Grassl Ass 66:119–126Google Scholar
  5. Bender A (1999) An impact of morphological and physiological transformations of red clover flowers accompanying polyploidization on the pollinators’ working speed and value as a guarantee for cross pollination. Agraarteadus 4:24–37Google Scholar
  6. Boelt B, Julier B, Karagić Đ, Hampton J (2015) Legume seed production meeting market requirements and economic impacts. Crit Rev Plant Sci 34:412–427CrossRefGoogle Scholar
  7. Boller B, Schubiger FX, Kölliker R (2010) Red clover. In: Boller B (ed) Handbook of plant breeding. Springer, Dordrecht, pp 439–455Google Scholar
  8. Bond DA (1968) Variation between tetraploid red clover plants in corolla tube length and height of nectar. J Agric Sci 71:113–116CrossRefGoogle Scholar
  9. Brodsgaard CJ, Hansen H (2002) Pollination of red clover in Denmark. DIAS Rep 71:1–50Google Scholar
  10. Büyükkartal HNB (2003) In vitro pollen germination and pollen tube characteristics in tetraploid red clover (Trifolium pratense L.). Turk J Bot 27:57–61Google Scholar
  11. Büyükkartal HNB (2008) Causes of low seed set in the natural tetraploid Trifolium pratense L. (Fabaceae). Afr J Biotechnol 7:1240–1249Google Scholar
  12. Büyükkartal HNB, Colgeçen HT (2007) The reasons of sterility during pollen grain formation in the natural tetraploid Trifolium pratense L. Int J Bot 3:188–195CrossRefGoogle Scholar
  13. Clifford PTP, Baird IJ (1993) Seed yield potential of white clover: characteristics, components and compromise. Proc XVII Int Grassl Congress 2:1678–1679Google Scholar
  14. Clifford PTP, Scott D (1989) Inflorescence, bumble bee and climate interactions in seed crops of a tetraploid red clover (Trifolium pratense L.). J Appl Seed Prod 7:38–45Google Scholar
  15. Davis AR (2001) Searching and breeding for structural features of flowers correlated with high nectar-carbohydrate production. Acta Hortic 561:107–121CrossRefGoogle Scholar
  16. Dennis BA (1980) Breeding for improved seed production in autotetraploid red clover. In: Hebbelethwaite PD (ed) seed production. Butterworths, London, pp 229–241Google Scholar
  17. Dijkstra J (1969) The importance of two-seeded pods of red clover (Trifolium pratense L.). Euphytica 18:340–351CrossRefGoogle Scholar
  18. Furuya H (2001) Comparisons of seed weight and seedling characteristics of diploid and autotetraploid red clover. Ph.D. thesis. University of Florida, FL, USAGoogle Scholar
  19. Grebenisan M, Savatti M (2011) The comparative effects of the micro- and macrosporogenesis of the red clover with different levels of ploidy, in relation with its fertility. Bull Univ Agric Sci Vet Med 68:138–143Google Scholar
  20. Guy P, Leconte D, Mousset-Declas C (1989) Les variétés de légumineuses de demain pour les régions atlantiques et continentales. Fourrages 119:281–297Google Scholar
  21. Hawkins RP (1969) Length of tongue in a honey bee in relation to the pollination of red clover. J Agric Sci 73:489–493CrossRefGoogle Scholar
  22. Herrmann D, Boller B, Studer B, Widmer F, Kölliker R (2006) QTL analysis of seed yield components in red clover (Trifolium pratense L.). Theor Appl Genet 112:536–545CrossRefPubMedGoogle Scholar
  23. Julén U (1950) Fertility conditions of tetraploid red clover. Hereditas 36:151–160CrossRefGoogle Scholar
  24. Julén G (1959) Rotklee, Trifolium pratense L. In: Parey P (ed) Handbuch der Pflanzenzüchtung. Band IV, Berlin, pp 239–305Google Scholar
  25. Kendal WA (1967) Growth of red clover pollen (II) Elongation in vitro. Crop Sci 7:342–344CrossRefGoogle Scholar
  26. Kouamé CN, Quesenberry KH (1993) Cluster analysis of a world collection of red clover germplasm. Genet Resour Crop Evol 40:39–47CrossRefGoogle Scholar
  27. Liatukas Ž, Bukauskaitė J (2012) Differences in yielding capability of diploid and tetraploid red clover in Lithuania. Proc Latv Acad Sci Sect B 66:163–167Google Scholar
  28. McGregor SE (1976) Insect pollination of cultivated crops. USDA Agricultural Research Service, Washington, DCGoogle Scholar
  29. Muntean L (2006) The variability of the morphological traits of tetraploid red clover cultivars studied in Cluj-Napoca environmental conditions. Not Bot Horti Agrobot Cluj Napoca 34:79–87Google Scholar
  30. Muntean L (2008) A comparative study of the variability of some morphological traits in a collection of diploid and tetraploid cultivars of red clover. In: Proceedings of the 43rd Croatian and 3rd International Symposium on Agriculture, pp 317–321Google Scholar
  31. Rao S, Stephen WP (2009) Bumble bee pollinators in red clover seed production. Crop Sci 49:2207–2214CrossRefGoogle Scholar
  32. Rumball W, Keogh RG, Miller JE (2003) “Crossway” and “Grasslands Broadway” red clovers (Trifolium pratense L.). New Zeal J Agr Res 46:57–59CrossRefGoogle Scholar
  33. Starling TM, Wilsie CP, Gilbert NW (1950) Corolla tube length studies in red clover. Agron J 42:1–8CrossRefGoogle Scholar
  34. Taylor NL (2008) A century of red clover breeding developments in the United States. Crop Sci 48:1–13CrossRefGoogle Scholar
  35. Taylor NL, Quesenberry KH (1996) Red clover science. Kluwer Academic Publishers, DordrechtCrossRefGoogle Scholar
  36. Van Minnebruggen A, Cnops G, Saracutu O, Goormachtig S, van Bockstaele E, Roldán-Ruiz I, Rohde A (2014) Processes underlying branching differences in fodder crops. Euphytica 195:301–313CrossRefGoogle Scholar
  37. Vleugels T, Cnops G, Roldán-Ruiz I (2014) Improving seed yield in red clover through marker-assisted parentage analysis. Euphytica 200:305–320CrossRefGoogle Scholar
  38. Vleugels T, Roldán-Ruiz I, Cnops G (2015) Influence of flower and flowering characteristics on seed yield in diploid and tetraploid red clover. Plant Breed 134:56–61.CrossRefGoogle Scholar
  39. Vleugels T, Roldán-Ruiz I, Ceuppens B, Smagghe G, Cnops G (2016) Are corolla tube dimensions the reason for low seed yield in tetraploid red clover? In: Roldán-Ruiz I et al. (eds.), Breeding in a world of scarcity: proceedings of the 2015 meeting of the section “Forage Crops and Turfgrasses” of Eucarpia. Springer, Dordrecht. doi:  10.1007/978-3-319-28932-8_44
  40. Woodfield DR, Baird IJ, Clifford PTP (2004) Genetic control of white clover seed yield potential. Proc New Zeal Grassl Ass 66:111–117Google Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2016

Authors and Affiliations

  • Tim Vleugels
    • 1
  • Bob Ceuppens
    • 1
    • 2
  • Gerda Cnops
    • 1
  • Peter Lootens
    • 1
  • Frederik R. D. van Parijs
    • 1
  • Guy Smagghe
    • 2
  • Isabel Roldán-Ruiz
    • 1
  1. 1.Plant Sciences UnitInstitute for Agricultural and Fisheries Research (ILVO)MelleBelgium
  2. 2.Department of Crop Protection, Faculty of Bioscience EngineeringGhent UniversityGhentBelgium

Personalised recommendations