Sains Malaysiana 44(7)(2015): 913–920

 

Thidiazuron Induces High Frequency Direct Somatic Embryogenesis Growth from Cotyledon Culture of Eurycoma longifolia

(Tidiazuron Merangsang Pertumbuhan Somatik Embriogenesis daripada Kultur Kotiledon Eurycoma longifolia)

 

ZAWAWI DHIYA DALILA*, JAÕAFAR HAFSAH, ZAINUDDIN ROKIAH,

KARI RODZIAH & MOHD NOOR MADIHAH

 

Faculty of Bioresources and Food Industry, Universiti Sultan Zainal Abidin, Kampus Tembila

22200 Besut, Terengganu Darul Iman, Malaysia

 

Diserahkan: 13 September 2014/Diterima: 5 Februari 2015

 

ABSTRACT

Eurycoma longifolia is one of the well-known herbal plants in Southeast Asian region due to its remarkable properties, especially the root part that can be used as aphrodisiac, anti-cancer, anti-malaria and anti-ulcer agent. Uncontrolled harvesting of this plant has reduced its population. Thus, an efficient protocol involving the induction of direct somatic embryogenesis from cotyledon culture of E. longifolia has been developed. 15% explants forming embryos were achieved on Modified Murashige and SkoogÕs (MMS) medium supplemented with 0.1 mg/L zeatin plus 0.2 mg/L indole-3-butyric acid (IBA). However, the addition of 0.12 mg/L thidiazuron (TDZ) has increased the percentage up to 22.0%. The maturation phase occurred in the same medium, thus decreased the frequency of subculture and the genetic instability of the embryos. Secondary somatic embryogenesis growth of 20%, which was the highest percentage of growth, was achieved from the primary embryos that were cultured in the MMS medium with the treatment of 2.0 mg/L IBA plus 0.075 mg/L TDZ. The results indicated that the direct somatic embryogenesis obtained can be further used to develop a protocol for plantlet regeneration of E. longifolia.

 

Keywords: Cotyledon; Eurycoma longifolia; secondary embryogenesis; thidiazuron; zeatin

 

ABSTRAK

Eurycoma longifolia adalah tumbuhan herba yang terkenal di kawasan Asia Tenggara memandangkan potensinya yang telah dikenal pasti terutamanya bahagian akar untuk mengubati malaria, ulser, kanser dan lemah tenaga batin. Penuaian tumbuhan ini yang dilakukan tanpa kawalan telah menyebabkan populasinya semakin berkurangan. Maka satu protokol cekap melibatkan aruhan secara terus somatik embriogenesis daripada pengkulturan kotiledon E. longifolia telah dijalankan. Pertumbuhan embrio sebanyak 15% telah berjaya dicapai menerusi media pertumbuhan Modified Murashige dan SkoogÕs (MMS) yang ditambah 0.1 mg/L zeatin dan 0.2 mg/L indole-3-butyric acid (IBA). Namun, penambahan 0.12 mg/L tidiazuron (TDZ) telah meningkatkan kadar pertumbuhan sehingga 22.0%. Pertumbuhan dan pertukaran embrio somatik ke fasa matang berlaku di dalam media pertumbuhan yang sama, oleh itu ia mengurangkan kekerapan subkultur dan ketidakstabilan genetik embrio tersebut. Pertumbuhan embrio sekunder sebanyak 20.0% iaitu pertumbuhan paling tinggi, telah dicapai daripada embroid primer yang dikulturkan di dalam media pertumbuhan MMS yang ditambah 2.0 mg/L IBA dan 0.075 mg/L TDZ. Keputusan ini menunjukkan pertumbuhan terus embrio somatik yang diperoleh berpotensi untuk diaplikasikan ke peringkat seterusnya dalam mikropropagasi E. longifolia.

 

Kata kunci: Embrio sekunder; Eurycoma longifolia; kotiledon; tidiazuron; zeatin

 

RUJUKAN

 

Anthony, J.M., Senaratna, T., Dixon, K.W. & Sivasithamparam K. 2004. Somatic embryogenesis for mass propagation of Ericaceae - A case study with Leucopogon verticillatus. Plant Cell Tissue Organ Culture 76(2): 137-146.

Balzon, T.A., Luis, Z.G. & Scherwinski-Pereira, J.E. 2013. New approaches to improve the efficiency of somatic embryogenesis in oil palm (Elaeis guineensis Jacq.) from mature zygotic embryos. In Vitro Cellular and Developmental Biology - Plant 49(1): 41-50.

Bell, L.M., Trigiano, R.N. & Conger, B.V. 1993. Relationship of abscisic acid to somatic embryogenesis in Dactylis glomerata. Environmental and Experimental Botany 33: 495-499.

Chan, K.L., Choo, C.Y., Abdullah, N.R. & Ismail, Z. 2004. Antiplasmodial studies of Eurycoma longifolia Jack using the lactate dehydrogenase assay of Plasmodium falciparum. Journal of Ethnopharmacology 92: 223-227.

Chan, K.L., OÕNeill, M.J., Phillipson, J.D. & Warhurst, D.C. 1986. Plants as sources of antimalarial drugs. Part 3: Eurycoma longifolia. Planta Medica 52: 105-107.

Choo, C.Y., Nah, B.K., Ibrahim, P. & Chan, K.L. 2000. Antimicrobial activity of Eurycoma longifolia Jack. In Interdisciplinary Approaches in Natural Products Research, edited by Rahmani, M., Mahmood, M., Sukari, M.A., Lian, G.E.C., Hin, T.Y.Y. & Ali, D.A.I. Proceedings of the 16th National Seminar on Natural Products Department.

Dhandapani, M., Kim, D.H. & Hong, S.B. 2007. Efficient plant regeneration via somatic embryogenesis and organogenesis from the explants of Catharanthus roseus. In Vitro Cellular and Developmental Biology - Plant 44(1): 18-25.

Fuentes, S.R.L., Calheiros, M.B.P., Manetti-Filho, J. & Vieira, L.G.E. 2000. The effects of silver nitrate and different carbohydrate sources on somatic embryogenesis in Coffea canephora. Plant Cell Tissue Organ Culture 60: 5-13.

Fujimura, T. & Komamine, A. 1975. Effects of various growth regulators on the embryogenesis in a carrot cell suspension culture. Plant Science Letters 5: 359-364.

Hamzah, S. & Yusof, A. 2003. The ergogenic effects of Eurycoma longifolia Jack: A pilot study. British Journal of Sports Medicine 37: 464-70.

Hussein, S., Ibrahim, R., Ling, A.P.K., Fadzillah, N.M. & Daud, S.K. 2005. Micropropagation of Eurycoma longifolia Jack via formation of somatic embryogenesis. Asian Journal of Plant Science 4: 472-485.

Ismail, Z., Ismail, N. & Lassa, J. 1999. Malaysian Herbal Monograph. Malaysian Monograph Committee, Kuala Lumpur.

JimŽnez, V.M. 2005. Involvement of plant hormones and plant growth regulators on in vitro somatic embryogenesis. Plant Growth Regulators 47: 2-3.

Kardono, L.B.S., Angerhofer, C.K., Tsauri, S., Padmawinata, K., Pezzuto, J.M. & Kinghorn, A.D. 1991. Cytotoxic and antimalarial constituents of the roots of Eurycoma longifolia. Journal of Natural Products 54: 1360-1367.

Klin, W., Suppl, W., Wernsdorfer, W.H., Ismail, S., Chan, K.L., Congpuong, K. & Wernsdorf, G. 2009. Activity of Eurycoma longifolia root extract against Plasmodium falciparum in vitro. The Middle European Journal of Medicine 121(3): 23-26.

Kuo, P.C., Shi, L.S., Damu, A.G., Su, C.R., Huang, C.H., Ke, C.H., Wu, J.B., Lin, A.J., Bastow, K.F., Lee, K.H. & Wu, T.S. 2003. Cytotoxic and antimalaria β- carboline alkaloids from the root of Eurycoma longifolia. Journal of Natural Products 66: 1324-1327.

Li, Z., Traore, A., Maximova, S. & Guiltinan, M.J. 1998. Somatic embryogenesis and plant regeneration from floral explants of cacao (Theobroma cacao L.) using thidiazuron. In Vitro Cellular and Developmental Biology - Plant 34: 293-299.

Murashige, T. & Skoog, F. 1962. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologia Plantarum 15: 473-497.

Naing, A.H., Kim, C.K., Yun, B.J., Jin, J.Y. & Lim, K.B. 2012. Primary and secondary somatic embryogenesis in Chrysanthemum cv. Euro. Plant Cell Tissue Organ Culture 112(3): 361-368.

Nishiwaki, M., Fujino, K., Koda, Y., Masuda, K. & Kikuta, Y. 2000. Somatic embryogenesis induced by the simple application of abscisic acid to carrot (Daucus carota L.) seedlings in culture. Planta 211: 756-759.

Pinto, G., Silva, S., Park, Y.S., Neves, L., Araœjo, C. & Santos, C. 2008. Factors influencing somatic embryogenesis induction in Eucalyptus globulus Labill.: basal medium and anti-browning agents. Plant Cell Tissue Organ Culture 95(1): 79-88.

Rai, V.R. & McComb, J. 2002. Direct somatic embryogenesis from mature embryos of sandalwood. Plant Cell Tissue Organ Culture 69: 65-70.

Raemakers, C.J., Jacobsen, E. & Visser, R.G. 1995. Secondary somatic embryogenesis and applications in plant breeding (review). Euphytica 81(1): 93-107.

Ramakrishnan, M. & Ceasar, S.A. 2013. Efficacious somatic embryogenesis and fertile plant recovery from shoot apex explants of onion (Allium cepa L.). In Vitro Cellular and Developmental Biology - Plant 49: 285-293.

Rashid, A. 2002. Somatic embryogenesis from immature and mature embryos of a minor millet Paspalum scrobiculatum L. Plant Cell Tissue Organ Culture 69: 71-77.

Rocha, D.I., Vieira, L.M., Tanaka, F.A.O., da Silva, L.C. & Otoni, W.C. 2012. Somatic embryogenesis of a wild passion fruit species Passiflora cincinnata Masters: Histocytological and histochemical evidences. Protoplasma 249(3): 747-58.

Rosli, N., Maziah, M., Chan, K.L. & Sreeramanan, S. 2009. Factors affecting the accumulation of 9-methoxycanthin- 6-one in callus cultures of Eurycoma longifolia. Journal of Forestry Research 20(1): 54-58.

Ruduś, I., Kępczyński, J. & Kępczyńska, E. 2001. The influence of the jasmonates and abscisic acid on callus growth and somatic embryogenesis in Medicago sativa L. tissue culture. Acta Physiologiae Plantarum 23(1): 103-107.

Santa-Catarina, C., Silveira, V., Balbuena, T.S., Viana, A.M., Estelita, M.E.M., Handro, W. & Floh, E.I.S. 2006. IAA, ABA, polyamines and free amino acids associated with zygotic embryo development of Ocotea catharinensis. Plant Growth Regulators 49(3): 237-247.

Shi, X., Dai, X., Liu, G., Zhang, J., Ning, G. & Bao, M. 2010. Cyclic secondary somatic embryogenesis and efficient plant regeneration in camphor tree (Cinnamomum camphora L.). In Vitro Cellular and Developmental Biology -Plant 46(2): 117-125.

Śliwińska, A., Olszowska, O., Furmanowa, M. & Nosov, A. 2008. Rapid multiplication of Polyscias filicifolia by secondary somatic embryogenesis. In Vitro Cellular and Developmental Biology - Plant 44(2): 69-77.

Stasolla, C. & Yeung, E.C. 2003. Recent advances in conifer somatic embryogenesis: Improving somatic. Plant Cell Tissue Organ Culture 74: 15-35.

Tee, T.T. & Azimahtol, H.L. 2005. Induction of apoptosis by Eurycoma longifolia Jack extracts. Anticancer Research 25: 2205-2213.

Vasic, D., Alibert, G. & Skoric, D. 2001. Protocols for efficient repetitive and secondary somatic embryogenesis in Helianthus maximiliani. Plant Cell Reports 20: 121-125.

Wu, H.C., Toit, E.S. & Reinhardt, C.F. 2007. A protocol for direct somatic embryogenesis of Protea cynaroides L. using zygotic embryos and cotyledon tissues. Plant Cell Tissue Organ Culture 89(2): 217-224.

Zakaria, Y., Rahmat, A., Hawariah, A., Pihie, L., Abdullah, N.R. & Houghton, P.J. 2009. Eurycomanone induce apoptosis in HepG2 cells via up-regulation of p53. Cancer Cell International 21: 1-21.

 

 

*Pengarang untuk surat-menyurat; email: ddalila@unisza.edu.my