Skip to main content
SpringerLink
Log in

Polyploidy Induction and Ploidy Level Determination in Annual and Perennial Diploid Medicago Species Using the Enumeration of Chloroplasts of Stomata Guard Cells

  • Published:
Cytology and Genetics Aims and scope Submit manuscript

Abstract

Primary screening of original diploid species and induced autotetraploids after induction of single node cutting of three populations of perennial diploid Medicago sativa ssp. caerulea: (Karaj1, Karaj2 and Tehran) and five annual diploid alfalfa species: (M. lupulina, M. radiata, M. rijidula, M. truncatula, M. turbinata) with three concentrations (0.10, 0.50, 1%) of colchicine counting the chloroplasts number in the stomata guard cells was done. In addition, the ploidy levels of species were determined using usual method of chromosomes counting in the metaphase cells of root tip meristems. Results of ANOVA showed significant effects of species, colchicine concentration and their interaction for survival and polyploidy induction rate (p < 0.01). The highest and lowest survival rate (80.2 and 8.9%) and tetraploidy induction (74.3 and 20.8%) were obtained in 0.10 and 1.0% colchicine concentration, respectively. In comparison, between species, the highest survival and tetraploidy induction rate were observed in M. truncatula (80.2 and 100%) followed by M. lupulina (73.6 and 73.6%), respectively. In contrast, the lowest survival and tetraploidy induction rate were observed in M. turbinata with 34 and 27.4%, respectively. Chloroplasts enumeration in stomata cell guards showed that in the survived species the number of chloroplasts of stomata guard cells were ranged between 8.07 to 9.15 in diploids and between 14.94 to 15.38 in tetraploids from induced diploids and five local tetraploid cultivated alfalfa cultivars, respectively. There was no significant difference between overall means of perennials and annual diploid alfalfa species for survival and tetraploidy induction. Ploidy levels have shown strong relationships with mean numbers of chloroplasts in stomata guard cells. It was concluded that enumeration of stomata guard cells, chloroplasts is an indirect and precise method for evaluating ploidy level in alfalfa.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1.
Fig. 2.

Similar content being viewed by others

REFERENCES

  1. Cardi, T., Carputo, D., and Frusciante, L., In vitro shoot regeneration and chromosome doubling in 2x and 3x potato clones, Am. Potato J., 1992, vol. 69, pp. 1–12. https://doi.org/10.1007/BF02853404

    Article  Google Scholar 

  2. Callum, J.B., Dixon, R.A., Farmer, A.D., et al., The Medicago Genome Initiative: a model legume database, Nucleic Acids Res., 2001, vol. 29, no. 1, pp. 114–117. https://doi.org/10.1093/nar/29.1.114

    Article  Google Scholar 

  3. Chen, L.L. and Gao, Sh.L., In vitro tetraploid induction and generation of tetraploids from mixoploids in Astragalus memberanaceus, Sci. Hortic. (Amsterdam), 2007, vol. 112, no. 3, pp. 339–334. https://doi.org/10.1016/j.scienta.2006.12.045

    Article  CAS  Google Scholar 

  4. Ghanavati, F. and Nematpajooh, N., Study of ploidy level of annual species of Onobrychis in Iran, Caryologia, 2012, vol. 65, no. 4, pp. 328–334. https://doi.org/10.1080/00087114.2012.760880

    Article  Google Scholar 

  5. Ghanavati, F., Mozafari, J., and Masumi, A.A., Determination of ploidy level with counting the chloroplast number in stomatal guard cells in Medicago sp., Seed Plant J., 2004, vol. 20, pp. 117–127.

    Google Scholar 

  6. Gu, X.F., Yang, A.F., Meng, H., and Zhang, J.R., In vitro induction of tetraploid plants from diploid Zizyphus jujuba Mill. cv. Zhanhua, Plant Cell Rep., 2005, vol. 24, pp. 671–676. https://doi.org/10.1007/s00299-005-0017-1

    Article  CAS  PubMed  Google Scholar 

  7. Han, D.S., Niimi, Y., and Nakamo, M., Production of doubled haploid plants through colchicine treatment of anther-derived haploid calli in the Asiatic hybrid lily 'Connecticut King’, 1999, J. Jpn. Soc. Hortic. Sci., vol. 68, pp. 979–983. https://doi.org/10.2503/jjshs.68.979

    Article  CAS  Google Scholar 

  8. Hawke, J.C. and Leech, R.M., Acetyl coenzyme A carboxylase in species of Triticum of different ploidy, Planta, 1990, vol. 181, no. 4, pp. 543–546. https://doi.org/10.1007/BF00193008

    Article  CAS  PubMed  Google Scholar 

  9. Koorneef, M., Van Diepen, J.A.M., Hanhart, C.J., et al., Chromosomal instability in cell- and tissue cultures of tomato haploids and diploids, Euphytica, 1989, vol. 43, pp. 179–186. https://doi.org/10.1007/BF00037911

    Article  Google Scholar 

  10. Lapiņa, L., Grauda, D., and Rashal, I., Characterization of Latvian alfalfa Medicago sativa genetic resources, Acta Biol. Univ. Daugavp., 2011, vol. 11, no. 2, pp. 134–140.

    Google Scholar 

  11. Leech, R.M., Observation of the mechanism of chloroplast division in higher plants, New Phytol., 1981, vol. 87, no. 1, pp. 1–9. https://doi.org/10.1111/j.1469-8137.1981.tb01686.x

    Article  Google Scholar 

  12. Malekzadeh Shafaroodi, S.A., Ghani, A.S., Habibi, M.P., and Amiri, A., Evaluation of polyploidy induction in basil (Ocimumbasilicum. L), J. Hortic. Sci., 2011, vol. 25, no. 4, pp. 461–469.

    Google Scholar 

  13. Melnychuk, O.V., Ozheredov, S.P., Rakhmetov, D.B., et al., The technology used for synthetic polyploid production of Miscanthus as cellulosic biofuel feedstock, Open Agric. J., 2020, vol. 14, pp. 164–173. https://doi.org/10.2174/1874331502014010164

    Article  CAS  Google Scholar 

  14. Melnychuk, O.V., Ozheredov, S.P., Rakhmetov, D.B., et al., Induction of polyploidy in giant miscanthus (Miscanthus × giganteus Greef et Deu.), Proc. Latv. Acad. Sci., Sect. B, 2020, vol. 74, no. 3, pp. 206–214. https://doi.org/10.2478/prolas-2020-0032

    Article  CAS  Google Scholar 

  15. Mirzai NeduShen, H., Annual Medics (Genetics and Breeding), Tehran: Res. Inst. For. Rangel., 2001.

    Google Scholar 

  16. Murray, H. and Standing, L., Genomic constancy during the development of Lathyrus odoratus cultivars, Heredity, 1992, vol. 68, pp. 321–327. https://doi.org/10.1038/hdy.1992.46

    Article  Google Scholar 

  17. Niazian, M. and Nalousi, A.M., Artificial polyploidy induction for improvement of ornamental and medicinal plants, Plant Cell, Tissue Organ Cult., 2020, vol. 142, no. 3, pp. 447–469. https://doi.org/10.1007/s11240-020-01888-1

    Article  Google Scholar 

  18. Omidbaigi, R., Mirzaee, M., Hassani, M.E., and Sedghi Moghadam, M., Induction and identification of polyploidy in basil (Ocimum basilicum L.) medicinal plant by colchicine treatment, 2020, Int. J. Plant Prod., vol. 4, no. 2, pp. 87–98. https://doi.org/10.22069/IJPP.2012.686

    Article  Google Scholar 

  19. Quan, K., Guolu, L., Qigao, G., and Xiaolin, L., Polyploid induction of Arctium lappa by colchicine, Plant Physiol. Commun., 2004, vol. 40, no. 2, pp. 157–158.

    Google Scholar 

  20. Sakiroglu, M., Sherman-Broyles, S., Story, A., Moore, K.J., Doyle, J.J., and Brummer, E.C., Patterns of linkage disequilibrium and association mapping in diploid alfalfa (M. sativa L.), Theor. Appl. Genet., 2012, vol. 125, no. 3, pp. 577–590. https://doi.org/10.1007/s00122-012-1854-2

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Sarathum, S., Hegele, M., Tantiviwat, S., and Nanakorn, M., Effect of concentration and duration of colchicine treatment on polyploidy induction in Dendrobium scabrilingue, L., Eur. J. Hortic. Sci., 2010, vol. 75, no. 3, pp. 123–127.

    CAS  Google Scholar 

  22. Singsit, C. and Ozias-Akins, P., Rapid estimation of ploidy levels of in vitro regenerated interspecific Arachis hybrids, Euphytica, 1992, vol. 64, pp. 183–188. https://doi.org/10.1007/BF00046047

    Article  Google Scholar 

  23. Sleper, D.A. and Poehlman, J.M., Breeding Field Crops, Oxford: Blackwell, 2006.

    Google Scholar 

  24. Su-Jin, B., Md Mazharul, I., Hong-Yul, K., and Ki-Byung, L., Induction of tetraploidy in watermelon with oryzalin treatments, Hortic. Sci. Technol., 2020, vol. 38, no. 3, pp. 385–393. https://doi.org/10.7235/HORT.20200037

    Article  Google Scholar 

  25. Touchell, D.H., Palmer, I.E., and Ranney, T.G., In vitro ploidy manipulation for crop improvement, Front. Plant Sci., 2020, vol. 11, art. ID 722. https://doi.org/10.3389/fpls.2020.00722

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Plant breeding, Science and Research Branch, Islamic Azad University, Tehran, Iran

    Elham Ansari & Mahmood Khosrowshahli

  2. Department of Plant breeding and Biotechnology, Kermanshah Branch, Islamic Azad University, Kermanshah, Iran

    Alireza Etminan

  3. Research Institute of Forests and Rangelands, Agricultural Research, Education and Extension Organization (AREEO), Tehran, Iran

    Ali Ashraf Jafari

Authors
  1. Elham Ansari
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mahmood Khosrowshahli
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Alireza Etminan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ali Ashraf Jafari
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Mahmood Khosrowshahli.

Ethics declarations

The authors declare that they have no conflicts of interest. This article does not contain any studies involving animals or human participants performed by any of the authors.

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ansari, E., Khosrowshahli, M., Etminan, A. et al. Polyploidy Induction and Ploidy Level Determination in Annual and Perennial Diploid Medicago Species Using the Enumeration of Chloroplasts of Stomata Guard Cells. Cytol. Genet. 56, 164–171 (2022). https://doi.org/10.3103/S0095452722020025

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.3103/S0095452722020025

Keywords:

Search

Navigation