Effects of different medium on morphophysiological traits of Lippia citriodora H.B.K. during in vitro culture condition

Document Type: Original article


1 Department of Horticulture, College of Agriculture, Tehran Science and Research branch, Islamic Azad University, Tehran, Iran;

2 Research Institute of Forests and Rangelands, Tehran, Iran;

3 National Institute of Genetic Engineering and Biotechnology, Tehran, Iran;

4 Department of Plant Sciences, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran;


Background & Aim: Lemon verbena (Lippia citriodora H.B.K.) is a richly scented herb and has a long history as a sacred and medicinal plant. Select an appropriate medium in tissue culture is a necessity for success Experimental: To evaluate the effect of 10 medium of previous researches for Lippia micropropagation (1- MS medium supplemented with BAP (3 mgL-1) + IBA (0.1 mgL-1) + activated charcoal (1 gL-1) 2- MS medium supplemented with BAP (1 mgL-1) + IBA (0.5 mgL-1) + activated charcoal (1 gL-1) 3- 1/2MS 4- MS medium supplemented with BAP (5 mgL-1) 5- MS medium supplemented with BAP (1 mgL-1) 6- MS medium supplemented with activated charcoal (1 gL-1) 7- MS medium supplemented with BAP (1 mgL-1) + activated charcoal (1 gL-1) 8- MS medium supplemented with BAP (1 mgL-1) + AgNO3 (5 mgL-1) 9- MS medium supplemented with AgNO3 (5 mgL-1) 10- MS (control)), an experiment was conducted as a Completely Randomized Design (CRD) with three replicates and three explants per replication.  Results & Discussion: The results showed positive effects for BAP (5 mgL-1) and somewhat active charcoal, but contrary to predictions, AgNO3 had no effect on reducing senescence and leaf necrosis. Recommended applications/industries: Therefore, Based on the finding, the four medium (MS medium supplemented with BAP (5 mgL-1)) with the highest rates of shoot fresh weight, leaf number, internodes number, shoot number, root fresh weight, callus diameter and root dry weight can be introduced as a suitable medium of tissue culture in this experimental conditions.


Article Title [Persian]

اثر محیط های مختلف کشت بر برخی صفات مورفوفیزیولوژیک به لیمو در شرایط کشت درون شیشه ای

Authors [Persian]

  • حسن نورافکن 1
  • فاطمه سفیدکن 2
  • سیدامیر موسوی 3
  • مظفر شریفی 4
  • احمد خلیقی 1
1 گروه علوم باغبانی، دانشکده کشاورزی، واحد علوم و تحقیقات، تهران، ایران؛
2 موسسه تحقیقات جنگل ها و مراتع کشور، تهران، ایران؛
3 پژوهشگاه ملی مهندسی ژنتیک و زیست فناوری، تهران، ایران
4 گروه علوم گیاهی، دانشکده علوم زیستی، دانشگاه تربیت مدرس، تهران، ایران؛
Abstract [Persian]

مقدمه و هدف: به­لیمو با نام علمی Lippia citriodora H.B.K سال­هاست به عنوان یک گیاه دارویی و معطر کشت شده و مورد مصرف قرار می‌گیرد. تکثیر جنسی به­لیمو به علت عدم جوانه­زنی بذر معمول نیست و تکثیر آن غیرجنسی انجام می­شود. انتخاب محیط کشت مناسب یکی از ضرورت­های موفقیت در کشت بافت است. روش تحقیق: به منظور ارزیابی اثرات 10 محیط کشت مورد استفاده در تحقیقات کشت بافت (1- MS حاوی BAP (3 mgL-1) + IBA (0.1 mgL-1) + زغال فعال (1 gL-1)، 2- MS حاوی BAP (1 mgL-1) +IBA (0.5 mgL-1) + زغال فعال (1 gL-1)، 3- 1/2MS، 4- MS حاوی BAP (5 mgL-1)، 5- MS حاوی BAP (1 mgL-1)، 6- MS حاوی زغال فعال (1 gL-1)، 7- MS حاوی BAP (1 mgL-1) + زغال فعال (1 gL-1)، 8- MS حاوی BAP (1 mgL-1) + نیترات نقره (5 mgL-1)، 9- MS حاوی نیترات نقره (5 mgL-1) و 10- شاهد (MS)) بر صفات مورفوفیزیولوژیک به‌لیمو، آزمایشی در قالب طرح کاملاً تصادفی با سه تکرار و سه ریزنمونه در هر تکرار انجام شد. نتایج و بحث: نتایج اثرات مثبت BAP (5 mgL-1) و تا حدودی زغال­فعال را بر بسیاری از شاخص­های رشد نشان داد ولی برخلاف انتظار، نیترات­نقره تاثیری روی کاهش پیری و نکروز برگ نشان نداد. توصیه کاربردی/ صنعتی: با توجه به یافته­ها می­توان محیط کشت چهارم (MS حاوی BAP (5 mgL-1)) را با داشتن بالاترین میزان از شاخص­های وزن تر شاخساره، تعداد برگ، تعداد میانگره، تعداد شاخه، وزن تر ریشه، قطر کالوس و وزن خشک ریشه به عنوان محیط مناسب کشت بافت در شرایط آزمایشی اشاره شده انتخاب و معرفی نمود.

Keywords [Persian]

  • به لیمو بنزیل آدنین کشت بافت گیاهان دارویی BAP
Bagheri, E. and Saffari. M. 2002. Principles of plant tissue culture. University of Ferdowsi Mashhad press. pp. 406. (In Persian).

Dehghan, A., Ebadi, M.T., Naghdi badi, H.A., Shahriyari, F., Azizi, M. and Asghari, GH. 2010. An overview of the techniques in the production of tropane alkaloids. Journal of Medicinal Plants., Year 9. 1(32): 149-164. (In Persian).

Firoozi, A. 2009. Suspension cultures of the herb milk thistle, Evaluation of silymarin production and use of somaclonal variation by ISSR. M.Sc. Thesis of Agricultural Biotechnology. Colleague of Agriculture. Tabriz University. Pp. 130. (In Persian).

Ghaemi, E., Khorshidi, D., Moradi, A., Seifi, A., Mazendrani, M. and Bazouri, M. 2006. The Effect of Ethanol Extract of Lemon verbena on the Skin Infection due to Staphylococcus aureus in an Animal Model. Iranian Journal of Medicinal and Aromatic Plants., 22(3): 242-249. (In Persian).

Hosseini hashemzadeh, H., Mohebalipour, N. and Nourafcan, H. 2014. Surface disinfection effect of different treatments on the growth of Stevia (Stevia rebaudiana Bert.) explants in vitro condition. First National Conference on Medicinal Plants and sustainable agriculture. Hamadan. Pp. 10. (In Persian).

Kheiri, H., Safi khani, K. and Barati, E. 2006. Planting method of yield of Lemon verbena in Hamadan. Proceedings of the National Conference on the Sustainable Development of Medicinal Plants. Research Institute of Forests and Rangelands press. Pp: 249-250. (In Persian).

Moaveni, P. 2010. Medicinal plants. Volume I. Islamic Azad University – Shahre Qods Branch press. Pp: 398. (In Persian).

Mohebalipour, N. 2012. Effect of growth regulators on Citral of Melissa ecotypes in vitro condition and evaluation of Genetic structure by ISSR markers. PhD thesis if plant breeding. Colleague of Agriculture. Tabriz University. Pp: 166. (In Persian).

Montazeri, F., Omidi, M. and Imani, N. 2011. Comparing various methods of seed and embryo culture and charcoal effect in optimizing in vitro culture of Galbanum (Ferula gummosa Boiss.). Iranian Journal of Medicinal and Aromatic Plants., 26 (4): 583-595. (In Persian).

Mozaffariyan, V. 1999. Dictionary of Iranian Plant Names. Second Edition. Farhang Moaser press. pp. 596. (In Persian).

Nouroozi, P. 2007. Direct regeneration from Hyacinth bulbs explants of two varieties (Atlantic and Carnegie) in vitro conditions. M.Sc. thesis. Horticultural Science department. University of Tehran. Pp: 111. (In Persian).

Oladzad, A., Qaderi, A., Naqdi badi, H.A. and Zare karizi, A.R. 2013. Rapid propagation of Lemon verbena (Lippia citriodora H.B.K.) by in vitro culture. Journal of Medicinal Plants., 11(2): 145-153. (In Persian).

Rezaei, M.B. and Kamkar, J. 2000. Investigation of chemical composition of lemon verbena essential oils. Pajoohesh and Sazandegi in agriculture and Horticulture., 14(2): 13-15. (In Persian)

Sabeti, H. 2001. Iranian Forests, trees and shrubs. Yazd university press. Pp. 806. (In Persian).

Shahhosseini, R. and Omidbeigi, R. 2002. Evaluation of essential oil content and composition of Lemon verbena under organic. Seventh Congress of Horticultural Sciences. Pp: 2517-2519. (In Persian).

Shahhosseini, R., Ghorbani, H., Saleh, R. and Omidbeigi, R. 2012. Quantitative and qualitative evaluation of essential oil of lemon verbena seeds. Journal of Plant Production research., 17(4): 91-96. (In Persian).

Shams Ardakani, M.R., Haji Akhoondi, E., Jamshidi, A.H. and Mohammad Rafiee, P. 2006. Production and maintenance of plant cell cultures and comparison metabolites produced in callus and whole plant. Journal of Medicinal Plants., Fifth year. Volume 17. Pp. 21-26. (In Persian).

Zargari, E. 1993. Medicinal plants. University of Tehran press. Fifth Edition. Volume 6. Pp. 711-714. (In Persian).

Kumar, V., Parvatam, G. and Ravishankar, G.A. 2009. AgNO3 - a potential regulator of ethylene activity and plant growth modulator. Electronic Journal of Biotechnology., 2(2): 1-15.

Yang, C. and Sung, F. 2002. Effect of gibberellic acid and dormancy-breaking chemicals on flower development of Rhododendron pulchrum sweet and R. scabrum Don. Scientia Horticulturae., 83: 331-337.

Portmann, E., Lopez Nigro, M.M., Reides, C.G., Liesuy, S., Ricco, R.A., Wagner M.L., Gurni, A.A. and Carballo, M.A. 2012. Aqueous extracts of Lippia turbinate and Aloysia citriodora (Verbenaceae): assessment of antioxidant capacity and DNA damage. International journal of toxicology., 00(0)1-11.

Severin, C., Bruzzese, D., Di Sapio, O., Gattuso, M. and Gattuso, S. 2006. Evaluation of in vitro behaviour of Aloysia citriodora Palau: histogical and chemical study. Molecular medicinal chemistry., Vol.11: 19-20.

Catalan, C.A.N. and de Lampasona, M.E.P. 2002. The chemistry of the genus Lippia (Verbenaceae). In S.E. Kintzios, (ed.) Oregano: The genera Origanum and Lippia, Taylor and Francis; London 127-149.

Gupta, S.K., Khanuja, S.P.S. and Kumar, S. 2001. In vitro micropropagation of Lippia alba. Current Science., 81(2): 210-206.

Tripathi, L. and Tripathi, J.N. 2003. Role of biotechnology in medicinal plants. Tropical Journal of Pharmaceutical Research., 2: 243-253.

Nalwade, S.M., and Tsay, H.S. 2004. In vitro propagation of some important Chinese medicinal plants and their sustainable usage. In Vitro Cellular & Developmental Biology - Plant., 40: 143-154.

Van Eck, J.M. and Kitto, S.L. 1990. Callus initiation and regeneration in Mentha. HortScience., 25(7): 804-806.

Braga, V.F., Mendes, G., Oliveira, R.T.R., Soares, C.Q.G., Resende, C.F., Pinto, L.C., Santana, R.D., Viccini, L.F., Raposo, N.R.B. and Peixoto, P.H.P. 2012. Micropropagation, antinociceptive and antioxidant activites of extracts of Verbena litoralis Kunth (Verbenaceae). Anaisda academia brasileria deciecias., 84(1):139-147.

Turhan, H. 2004. The effect of silver nitrate (ethylene inhibitor) on in vitro shoot development in potato (Solanum tuberosum L.). Biotechnology., 3(1): 72-74.