نقش اکوتیپ، ریزنمونه و تنظیم‌کننده‌های رشد گیاهی در کشت تعلیقی سلولی چویل (Ferulago angulata (Schlecht.) Boiss.)

نوع مقاله : مقالات پژوهشی

نویسندگان

• سید مهران علوی ¹
• اسد معصومی اصل ²
• ناصر زارع ¹
• رسول اصغری زکریا ¹
• پریسا شیخ زاده مصدق ¹

¹ دانشگاه محقق اردبیلی

² دانشگاه یاسوج

https://doi.org/10.22067/jhorts4.v33i3.79173

چکیده

گیاه چویل دارای خواص داروئی مختلف بوده و تاکنون گزارشی از کشت تعلیقی سلولی این گیاه بومی ایران ارائه نشده است. به ‌منظور ارزیابی تأثیر عوامل مختلف بر پینه‌زایی چویل، آزمایشی به صورت فاکتوریل بر پایه طرح کاملاً تصادفی اجرا شد. فاکتورها شامل اکوتیپ (آب­نهر، گایونه، وزگ و سی‌سخت)، ریز نمونه (ریشه، برگ و ساقه) به همراه ترکیبات هورمونی مختلف NAA و BAP (هر کدام در غلظت‌های 0، 5/0، 1 و 2 میلی‌گرم در لیتر) در محیط کشت پایه موراشیگ و اسکوگ بودند. نتایج نشان دادند که بهترین اکوتیپ در واکنش به پینه‌زایی، اکوتیپ آب‌نهر و بهترین ریز‌نمونه، ریزنمونه برگی و بهترین ترکیب هورمونی شامل 1 میلی‌گرم در لیتر NAA و 5/0 میلی‌گرم در لیتر BAP بود. به‌ منظور بررسی تأثیر عوامل مختلف بر کشت تعلیقی سلولی چویل، علاوه بر هورمون NAA، از سطوح مختلف (غلظت‌های 0، 5/0، 1 و 2 میلی‌گرم در لیتر) هورمون 2,4-D نیز در ترکیب با هورمون BAP استفاده شد. بهترین ترکیب هورمونی برای کشت تعلیق سلولی، 2 میلی‌گرم بر لیتر NAA یا 2,4-D به همراه 5/0 میلی‌گرم بر لیتر BAP بود و بهترین ریزنمونه و اکوتیپ به ترتیب، ریزنمونه‌ برگی و اکوتیپ آب‌نهر بود. در مجموع، اکوتیپ آب نهر و ریزنمونه برگی هم در کشت پینه و هم در کشت تعلیقی سلولی نسبت به بقیه اکوتیپ­ها و ریزنمونه­ها برتری نشان دادند و بهترین ترکیب هورمونی در هر دو نوع کشت 2-1 میلی‌گرم در لیتر NAA و 5/0 میلی‌گرم در لیتر BAP بود.
 

کلیدواژه‌ها

• پینه
• تنظیم‌کننده رشد
• ژنوتیپ
• SCV
• PCV

20.1001.1.20084730.1398.33.3.13.3

عنوان مقاله [English]

The Role of Ecotype, Explant and Plant Growth Regulators on Cell Suspension Culture of Ferulago angulata (Schlecht.) Boiss.

نویسندگان [English]

• Seyyed Mehran Alavi ¹
• Asad Masoumiasl ²
• Naser Zare ¹
• Rasul Asghari Zakaria ¹
• Parisa Sheikhzade Mosaddegh ¹

¹ Mohaghegh Ardabili University

² Yasouj University

چکیده [English]

Introduction: The main habitat of Chavil, Ferulago angulata, in Iran is Zagros area. This plant has a rejuvenating effect and is used to treat digestive diseases and intestinal worms. Because the different explants show different amounts of callogenesis under the effect of different growth regulators, selection of an optimal explant and suitable plant growth regulators combination has a significant effect on the production of callus and their suspension culture. There is no reports on Ferulago angulata callogenesis and its cell suspension culture. Therefore, this study was designed and implemented to optimize callus production and cell suspension culture in this important medicinal plant.
Materials and Methods: Seeds of Chavil were collected from four different habitats in Kohgilooyeh and BoyerAhmad Province in Southwest of Iran include Abenahr, Guayoune, Vezg and Sisakhat. Seedlings obtained from embryo culture were used to prepare the explants. Various explants (leaf, root and stem) were cultured on MS medium supplemented with different concentrations (0, 0.5, 1 and 2 mgl-1) of NAA and BAP. Callus traits were evaluated and from the best culture medium, the best explants and the best PGRs composition for callogenesis of each ecotype were used to cell suspension culture. In order to study the growth rate of cells in suspension culture and plotting the curve of cells growth, two cell density indices and packed cell volume index were evaluated. To determine the cell density index, every 3 days, 10 ml of cell suspension were transferred to the graded falcon and centrifuged at 5000 g for 5 minutes, and the percentage of sediment cells was calculated as the total volume. To determine the packed cell volume index, also every 3 days, 10 ml of culture medium containing cells were transferred to the graded falcon and stored for 30 minutes to precipitate cells and cell masses. Finally, the cell volume was recorded and was calculated as percentage of the total collected medium.
Results and Discussion: According to the callogenesis percentage, the best ecotype is Abenahr and best explant is leaf explant. The highest level of NAA is 2 mgl-1, and the best level of BAP is 2 mgl-1, which causes 100 callogenesis percentage. The best medium for cell suspension culture is MS medium containing 2 mgl-1 NAA and 0.5 mgl-1 BAP for callus was obtained from leaf explant of Abenahr ecotype. Along with these plant growth regulators, 2,4-D was used in combination with BAP to form suspension culture. The results also showed that 2 mgl-1 2,4-D plus 0.5 mgl-1 BAP were useful in producing suspensions. The difference between 2,4-D +BAP and NAA + BAP combinations more cell volumes were observed, and cell suspension was created at a faster rate and in less time, which is an advantage in research work. Growth rate of cell suspension originated from the leaf explant was higher than root explant. In terms of culturing cell suspension, the Abenahr ecotype was favorable compared to other ecotypes. During cell suppression culture of Cyperus aromaticus by applying different levels of NAA, cell growth was increased up to 3 weeks after application, and then decreased. By applying 2,4-D, cell growth also increased until the third week, and after the third week, cell growth declined, which was very low growth rate compared with the NAA. In cell suspension culture of sugar beet, using 2,4-D was much more effective than NAA on all explants. In the present study, 2,4-D was also more effective than NAA for cell suspension culture of Chavil.
Conclusion: In general, the Abenahr was the best ecotype among of investigated. The explants in both callus culture and the suspension culture, and the best combination of plant growth regulator in both culture was 2 mgl-1 NAA plus 0.5 mgl-1 BAP.

کلیدواژه‌ها [English]

• Callus
• Genotype
• PCV
• Plant growth regulator
• SCV

مراجع

1-Abrazeh M. 2004. Investigation of some ecological characteristics of Chavil (Ferulago angulata) in protected Dena area. Ms.C thesis in Natural resources, Tarbiat Modarres University, Tehran. (In Persian)

2-Bhojwani S.S., and Razdan M.K. 1992. Plant tissue culture, Theory and practice. Elsevier, Amsterdam, London, New York, Tokyo.

3-Daud Z., and Keng C.L. 2006. Effects of plant growth regulators on the biomass of embryogenic cells of Cyperus aromaticus (Ridly) Mattf and Kukenth. Biotechnology 5:75-78.

4-Darderafshi M.J., Bahrami G.h., Sadeghi E., Khanahmadi M., Mohammadi M., and Mohammadi R. 2014. The effect of Ferulago angulata essential oil on Staphylococcus aureus during the manufacture and preservation of Iranian white cheese. Iranian Journal of Nutrition Sciences and Food Technology 8(4):13-20. (In Persian with English abstract)

5-Dixon R.A., and Gonzales R.A. 1996. Plant Cell Culture: A Practical Approach. 2nd Ed. IRL Press.

6-Ehsanpour A.A., and Amini F. 2003. Plant Cell and Tissue Culture. Jahad-Daneshgahi-Esfahan Publisher. (In Persian)

7-Evans D.E., Coleman J.O.D., and Kearns A. 2003. Plant cell culture: The basics. London, Taylor & Francis Publisher.

8-Farjaminezhad R., Zare N., Asghari-Zakaria R., and Farjaminezhad M. 2014. Establishment and optimization of cell growth in suspension culture of Papaver bracteatum a biotechnology approach for thebaine production. Turkish Journal of Biology 37:689-697.

9-Farsi A.R., and Zulali J. 2014. Principles of plant biotechnology. Six Edition. Mashhad Ferdousi University Press. Iran. (In Persian)

10-Foladvand Z., Fazelinasab B., darikvand R., and Gasmi-pirbaloti A. 2014. Optimization of Callus Induction and Cell Suspension in Catharanthus roseus. Journal of Herbal Drugs, 5(3):157-163. (In Persian)

11-Ghasemian K.h., Nazeri S., Chehregani-Rad A., and Mirzaie-Asl A. 2012. The stages of somatic embryogenesis derived from zygotic embryo of Dorema ammoniacum D. Journal of Cell & Tissue (JCT) 3(1): 21-27. (In Persian with English abstract)

12-Gurel S., Gurel E., and Kaya Z. 2001. Callus development and indirect shoot regeneration from seedling of sugar beet (Beta vulgaris L.) cultured in vitro. Turkish Journal of Botany 25: 25-33.

13-Hren M., Baebler S., Camloh M., Kovac M., Ravnikar M., and Zel J. 2006. Yew (Taxus x media Rehd.) cell suspension cultures as a source of taxanes. Acta Physiological Plantarum 28: 3-8.

14-Hussain Z., Haroon M., Bano R., Rashid H., and Chaudhry Z. 2010. Protocol optimization for efficient callus induction and regeneration in three Pakistani rice cultivars. Pakistan Journal of Botany, 42:879-887.

15-Janarthanam B., Gopalakrishnan M., Lakshmi Sai G., and Sekar T. 2009. Plant regeneration from leaf derived callus of Stevia rebaudiana Bertoni. Plant Tissue Culture and Biotechnology 19(2): 133-141.

16-Khalil S.M., and Elbanna A.A.M. 2003. Highly efficient somatic embryogenesis and plant regeneration via suspension cultures of banana (Musa spp.). Arab Journal of Biotechnology 7: 99-110.

17-Khawar K.M., Sarihan E.O., Sevimay C.S., Cocu S., Parmaksiz I., Uranbey S., Ipek A., Kaya M.D., Sancak C., and Özcan S. 2005. Adventitious shoot regeneration and Micropropagation of Plantago lanceolata L. Periodicum Biologorum 107:57–61.

18-Koohi L., Zare N., Asgari-zakaria R., Sheikhzadeh-Mosaddeg P., and Daryani P. 2013. The role of plant growth regulators and different explants on the tissue culture response and cell suspension in German chamomile (Matricaria chamomilla L.). Journal of Crop Ecophysiology 2 (30): 203-214. (In Persian with English abstract)

19-Kwon T.H., Abe T., and Sasahara T. 1993. Efficient callus induction and plant regeneration in Sesamum species. Plant Tissue and Culture Letters 10: 260-266.

20-Lee E., Mobin M., Hahn E.J., and Paek K.Y. 2006. Effects of sucrose, inoculum density, auxins, and aeration volume on cell growth of Gymnema sylvestre. Journal of Plant Biology 49: 427-431.

21-Lukmanul Hakkim F., Kalyani S., Essaa M., Girija S., and Song H. 2011. Production of rosmarinic acid in Ocimum sanctum (L.) cell suspension cultures by the influence of growth regulators. International Journal of Biological and Medical Research 2(4): 1158-1161.

22-Molina S.M. 2004. In vitro callus induction and plants from stem and petiole explants of Salvia canariensis L. Plant and Tissue Culture 14: 167-172.

23-Mortazavi R., Dehdari M., and Masoumiasl A. 2016. Study of Callus Induction of Medicinal Chavil Plant (Ferulago angulata B.) Using Types of Explants and Growth Regulators. Agricultural Biotechnology 14(2): 73-80. (In Persian with English abstract)

24-Mozaffarian V. 2007. Iranian Plant Culture Names. First Volume, First edition, Farhang-e- Moaser Publisher. Tehran, Iran. (In Persian)

25-Mujib A., Bandhyopadhyay S., and Ghosh P.D. 2000. Tissue culture derived plantlet variation in Caladium an important ornamental. Plant Cell, Tissue and Organ Culture 10: 149-155.

26-Murashige T., and Skoog F. 1962. A revised medium for rapid growth and bioassay with tobacco tissue cultures. Physiological Plantarum 15: 473-497.

27-Mythili J.B., and Thomas P. 1999. Micropropagation of Pointed Gourd (Trichosanthes dioica Roxb.). Scientia Horticulture 79: 87-90.

28-Ozgen M., Turet M., Altinok S., and Sancak C. 1998. Efficient callus induction and plant regeneration from mature embryo culture of winter wheat (Triticum aestivum L.) genotypes. Plant Cell Reports 18: 331-335.

29-Palai S.K., Rout G.R., and Das P. 1997. Micropropagation of ginger (Zingiber officinale Rosc.): interaction of growth regulators and culture conditions. Proceeding of Biotechnology of Spices, Medicinal and Aromatic Plants, Kerala, 20–24.

30-Perez-Bermudez P., Cornejo M.J., and Segura J. 1983. In vitro propagation of Digitalis obscura L. Plant Science Letters 30: 77-82.

31-Phillips G.C., Hubstenberger J.F., and Hansen E.E. 1995. Plant regeneration by organogenesis from callus and cell suspension cultures. Plant Cell, Tissue and Organ Culture, Heidelberg: Springer and Verlag, 67-78.

32-Ramirez-Malagon R., Borodanenco A., Barrera-Guerra J.L., and Ochoa-Alejo N. 2001. Shoot number and shoot size as affected by growth regulators in in vitro Cultures of Spathiphyllum floribundum L. Scientia Horticulture 89: 227-236.

33-Richard D., Lescot M., Inze D., and De Veylder L. 2002. Effect of auxin, cytokinin, and sucrose on cell cycle gene expression in Arabidopsis thaliana cell suspension cultures. Plant Cell, Tissue and Organ Culture 69: 167-176.

34-Senoussi M.M., Nora B., and Joe C. 2009. Impact of hypoxia on the growth and alkaloid accumulation in Catharanthus roseus cell suspension. Acta Physiologiae Plantarum 31: 359–362.

35. Sarabadani-tafreshi R., Omidi M., Bihamta M.R., and Davazdahemami S. 2008. Study of in vitro embryo culture and Effect of explants, different hormonal levels on callus induction and shooting of Ferula gummosa. Journal of Medicinal Plants 27: 71-81. (In Persian)

36-Shahadati-Moghadam Z. 2001. Plant growth regulators. M.Sc. Thesis of agriculture, Mazandaran University, Iran. (In Persian)

37-Sorbi E., Moradi a., Masoumiasl A., and Balochi H.R. 2017. Optimization of Callus Induction and somatic embryogenesis in twoGenotypes of medicinal Chavil plant (Ferulago angulata L.). Plant Production Research 21(3): 41-62. (In Persian)

38-Silveira V., Floh E.I.S., Handro W., and Pedro Guerra M. 2004. Effect of plant growth regulators on the cellular growth and levels of intracellular protein, starch and polyamines in embryogenic suspension cultures of Pinus taeda. Plant Cell, Tissue and Organ Culture 76: 53-60.

39-Street H.E. 1977. Plant tissue and cell culture. 2nd Edition, Blackwell. Oxford.

40-Taha H.S., El-Bahr M.K., and Seif-El-Nasr M.M. 2008. In vitro studies on Egyptian Catharanthus roseus (L.) G. Don.: 1- calli Production, direct shootlets Regeneration and alkaloids determination. Journal of Applied Science Research 4: 1017-1022.

41-Taran M., Ghasempour H.R., and Shirinpour E. 2010. Antimicrobial activity of essential oils of Ferulago angulata subsp. Carduchorum. Jundishapur Journal of Microbiology 3: 10-4. (In Persian)

42-Zare A., Solouki R., Omidi M., Irvani M., Mahdi-Nezad N., and Rezazadeh S.H. 2010. Callus induction and plant regeneration in Ferula assa-foetida L. (Asafetida), an endangered medicinal plant. Trakia Journal of Sciences 8(1): 11-18.