Micropropagation and molecular characterization of Thymus sibthorpii Benth. (Lamiaceae), an aromatic-medicinal thyme with ornamental value and conservation concern

  • Georgios TsoktouridisEmail author
  • Nikos Krigas
  • Virginia Sarropoulou
  • S. Kampouropoulou
  • Katerina Papanastasi
  • Katerina Grigoriadou
  • Georgios Menexes
  • Eleni Maloupa


Thymus sibthorpii Benth. (Lamiaceae), with accession number 01,1796-22, is a biotype of native Greek thyme with ascending stems and potential use as a new medicinal-aromatic crop and ornamental plant. An efficient and reliable protocol for in vitro clonal propagation of T. sibthorpii from nodes and meristem tip explants was developed. Shoot proliferation succeeded on a new basal medium (BB) without plant growth regulators, as prior experiments with 6-benzyladenine generated hyperhydricity. Eight different basal media were compared; on two formulations using the new BB 5.9 and 5.6 shoots per explant were produced. Regenerated single shoots were rooted in the BB medium, supplemented with 5 μM of indole-3-butyric acid, and produced 3.1 roots along with 2.5 adventitious shoots. Three types of acclimatization were assessed: in vitro, using two different systems (no significant differences); ex vitro, using eight soil substrates under greenhouse and outdoor nursery conditions (in two of them, 100% of plantlets survived); and in field cultivations, established at eight geographically distant areas of Greece (100% survival rate at all locations). Molecular characterization of T. sibthorpii was evaluated with one nuclear ribosomal DNA and seven chloroplast DNA markers, followed by DNA sequence comparisons with a total of 30 different Thymus species, subspecies, and varieties. The trnH/psbA, trnL/trnF, and matK genes were the most efficient markers for molecular characterization of T. sibthorpii. The molecular markers rpoC1 and petB/petD did not match to any Thymus species and therefore, these DNA sequences provide new sequence information for entire Thymus taxa.


Ascending thyme DNA barcoding In vitro clonal propagation Molecular markers Soil substrates and acclimatization 


Funding information

This research project was funded under the Project ‘Research & Technology Development Innovation Projects’-AgroETAK, MIS 453350, in the framework of the Operational Program ‘Human Resources Development’. It was co-funded by the European Social Fund through the National Strategic Reference Framework (Research Funding Program 2007-2013) coordinated by the Hellenic Agricultural Organization – DEMETER (Institute of Plant Breeding and Genetic Resources).


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Copyright information

© The Society for In Vitro Biology 2019

Authors and Affiliations

  • Georgios Tsoktouridis
    • 1
    Email author
  • Nikos Krigas
    • 1
  • Virginia Sarropoulou
    • 1
  • S. Kampouropoulou
    • 2
  • Katerina Papanastasi
    • 1
  • Katerina Grigoriadou
    • 1
  • Georgios Menexes
    • 2
  • Eleni Maloupa
    • 1
  1. 1.Laboratory of Conservation and Evaluation of Native and Floricultural Species-Balkan Botanic Garden of KroussiaInstitute of Plant Breeding and Genetic Resources, Hellenic Agricultural Organization – DemeterThermiGreece
  2. 2.Laboratory of Agronomy, School of AgricultureAristotle UniversityThessalonikiGreece

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