Effects of Plant Growth Regulators on Micropropagation of Black Pepper (Piper nigrum) in Commercial-Scales

Nguyen Ba Dat

School of Biotechnology, International University – Vietnam National University HCMC, Vietnam.

Tran Thi Hoang Anh

Western Highlands Agriculture and Forestry Science Institute, Vietnam.

Bui Thanh Hoa

School of Biotechnology, International University – Vietnam National University HCMC, Vietnam.

Pham Hong Diep

School of Biotechnology, International University – Vietnam National University HCMC, Vietnam.

Tran Van Minh *

School of Biotechnology, International University – Vietnam National University HCMC, Vietnam.

*Author to whom correspondence should be addressed.


Aim: Black pepper (Piper nigrum) is renowned as a popular spice used to flavor dishes around the world and is also an agricultural export commodity that brings high economic value to many countries. However, traditional propagation methods by seed, cuttings, layering, and grafting reveal many limitations such as the low multiplication coefficient, the low survival rate of seedlings, and especially when there is an outbreak of disease, the ability to control the disease source is exceedingly difficult, leading to the spread of disease. Therefore, an alternative method through apical meristem micropropagation has been applied to contribute to sustainable and disease free pepper cultivars. Propagation by apical shoot meristem gives high propagation efficiency, good and uniform seedling quality, and can be propagated in large numbers in a brief period.

Methodology: This study is conducted to contribute to optimizing the process of micropropagation of pepper plants to meet the needs of producing excellent quality seedlings, serving the Vietnamese pepper industry. The two steps in the micropropagation sequence studied in this project are shoot formation and bud cluster multiplication. In this study, we investigate the effects of BA (6-Benzylaminopurine) and NAA (α-Naphthalene acetic acid) in shoot formation. Regenerated shoots were then cultured in MS media supplemented with different combinations of Kinetin (6-Furfurylaminopurine) and NAA in bud cluster multiplication experiment.

Results: The results showed that the apical shoot meristem cultured on MS medium (Murashige and Skoog, 1962) supplemented with 0.5 mg L-1 NAA and 2.0 mg L-1 BA was the optimal medium for shoot formation rate (100%). Besides, MS culture medium supplemented with Kinetin 1.5 mg L-1 + NAA 0.5 mg L-1 produced superior results in bud cluster multiplication compared with other treatments (8, 20 buds) after 8 weeks (about 2 months).

Conclusion: BA, Kinetin and NAA give important rules on micropropagation of black pepper in commercial-scale.

Keywords: Black pepper, bud cluster, micropropagation, shoot formation, tissue culture

How to Cite

Dat, Nguyen Ba, Tran Thi Hoang Anh, Bui Thanh Hoa, Pham Hong Diep, and Tran Van Minh. 2024. “Effects of Plant Growth Regulators on Micropropagation of Black Pepper (Piper Nigrum) in Commercial-Scales”. Asian Journal of Biotechnology and Bioresource Technology 10 (2):45-59. https://doi.org/10.9734/ajb2t/2024/v10i2204.


Download data is not yet available.


Mathew PJ, Mathew PM, Kumar V. Graph clustering of Piper nigrum L. (black pepper). Euphytica. 2001;118(3):257–264. Available:https://doi.org/10.1023/A:1017554501828

Srinivasan K. Black pepper and its pungent principle-piperine: A review of diverse physiological effects. Critical Reviews in Food Science and Nutrition. 2007a;47(8): 735–748.

Shityakov S, Bigdelian E, Hussein AA, Hussain MB, Tripathi YC, Khan MU, Shariati MA. Phytochemical and pharmacological attributes of piperine: A bioactive ingredient of black pepper. European Journal of Medicinal Chemistry. 2019;176:149–161. Available:https://doi.org/10.1016/j.ejmech.2019.04.002

Babu KN, Lukose R, Ravindran PN. Tissue culture of tropical spices. Genetic Engineering, Molecular Biology and Tissue Culture for Crop Pest and Disease Management. 1993:257–267.

Du TX, Trang, NTH, Loan NTK, Tuấn TT, Phòng HT, Hà TTT, Giáp ĐĐ. Ứng dụng kỹ thuật nuôi cấy đỉnh sinh trưởng và lớp mỏng tế bào trong vi nhân giống cây Hồ tiêu giống Vĩnh Linh. Trong: Hội nghị khoa học công nghệ sinh học toàn quốc năm; 2013a.

Phượng TTB, Tuấn NĐ, Giang HTC. Nghiên cứu hệ thống tái sinh cây hồ tiêu (Piper nigrum). Hue University Journal of Science (HU JOS). 2014;94(6), Article 6. Available:https://jos.hueuni.edu.vn/index.php/TCKHDHH/article/view/1613

Ahmad N, Fazal H, Abbasi BH, Rashid M, Mahmood T, Fatima N. Efficient regeneration and antioxidant potential in regenerated tissues of Piper nigrum L. Plant Cell, Tissue and Organ Culture (PCTOC), 2010;102(1):129– 134. Available:https://doi.org/10.1007/s11240-010-9712-x

Atal CK, Banga SS. Phytochemical studies on stem of P. longum. Indian J. Pharm. 1962;24:105.

Hu CY, Wang PJ. Meristem, shoot tip and bud cultures. Handbook of Plant Cell Culture (USA); 1983 Available:https://scholar.google.com/scholar_lookup?title=Meristem%2C+shoot+tip+ and+bud+cultures&author=Hu%2C+C.Y.&publication_year=1983

Vinocur B, Carmi T, Altman A, Ziv M. Enhanced bud regeneration in aspen (Populus tremula L.) roots cultured in liquid media. Plant Cell Reports. 2000;19(12): 1146–1154. Availablehttps://doi.org/10.1007/s002990000243

Bhat SR, Chandel KPS, Malik SK. Plant regeneration from various expiants of cultivated Piper species. Plant Cell Reports. 1995;14(6):398– 402. Available:https://doi.org/10.1007/BF00238605

Sajc L, Grubisic D, Vunjak-Novakovic G. Bioreactors for plant engineering: An outlook for further research. Biochemical Engineering Journal. 2000;4(2):89–99. Available:https://doi.org/10.1016/S1369-703X(99)00035-2

Joseph B, Joseph D, Philip VJ. Plant regeneration from somatic embryos in black pepper. Plant Cell, Tissue and Organ Culture. 1996;47(1):87–90. Available:https://doi.org/10.1007/BF02318970

Nazeem PA, Joseph L, Geetha CK, Sreekandan Nair G. In vitro techniques for cloning of black pepper, Piper nigrum L. J. Plantation Crops. 1992;20:257–257.

Philip VJ, Joseph D, Triggs GS, Dickinson NM. Micropropagation of black pepper (Piper nigrum Linn.) through shoot tip cultures. Plant Cell Reports. 1992;12 (1):41–44. Available:https://doi.org/10.1007/BF00232421

Sujatha R, Babu LC, Nazeem PA. Histology of organogenesis from callus cultures of black pepper (Piper nigrum L.). Journal of Tropical Agriculture. 2006;41(0), Article 0.

Mai NT, Ngọc NTT, Anh TTH, Tân TV, Loan CTP, Thủy NTT. Xây dựng quy trình nhân giống cây hồ tiêu sạch bệnh bằng kỹ thuật nuôi cấy mô tế bào; 2020.

Murashige T, Skoog F. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologia Plantarum. 1962;15(3):473–497.

Cronauer SS, Krikorian AD. Multiplication of Musa from Excised Stem Tips. Annals of Botany, 1984;53(3):321–328. Available:https://doi.org/10.1093/oxfordjournals.aob.a086696

D’Agostino IB, Kieber JJ. Molecular mechanisms of cytokinin action. Current Opinion in Plant Biology. 1999;2(5):359–364. Available:https://doi.org/10.1016/S1369-5266(99)00005-9

Legesse T, Mekbib F, Gebre E. Effect of cytokinins concentration and types for shoot induction and multiplication experiments of In Vitro Propagation of Black Pepper (Piper nigrum L.) from Nodal Culture. International J. Agriculture Sciences. 2022;4:2348–3997.

Hussain A, Naz S, Nazir H, Shinwari Z. Tissue culture of black pepper (Piper nigrum L.) in Pakistan. Pak. J. Bot. 2011; 43:1069–1078.

Zhao D, Hu G, Chen Z, Shi Y, Zheng L, Tang A, Long C. Micropropagation and in vitro flowering of Dendrobium wangliangii: A critically endangered medicinal orchid. J Med Plants Res. 2013;7(28): 2098–2110

Gaspar Th, Kevers C, Faivre-Rampant O, Crèvecoeur M, Penel CL, Greppin H, Dommes J. Changing concepts in plant hormone action. In Vitro Cellular & Developmental Biology - Plant, 2003; 39(2):85–106. Available:https://doi.org/10.1079/IVP2002393

Thimann KV. Auxins and the Inhibition of Plant Growth. Biological Reviews. 1939;14(3):314–337. Available:https://doi.org/10.1111/j.1469185X.1939.tb00937.x

Cleland RE. Auxin and Cell Elongation. In P. J. Davies (Ed.), Plant Hormones and their Role in Plant Growth and Development. Springer Netherlands. 1987: 132–148. Available:https://doi.org/10.1007/978-94-009-3585-3_8

Kieber JJ, Schaller GE. Cytokinins. The Arabidopsis Book / American Society of Plant Biologists. 2014;12:e0168. Available:https://doi.org/10.1199/tab.0168

Thuyen DA, Du, TX, Giap DD, Ton NT. Preliminary study on the micropropagation in vitro of black pepper (Piper nigrum L.). Academia Journal of Biology2005;27(3) Article 3. Available:https://doi.org/10.15625/08667160/v27n3.5268