Culture conditions for induction of hairy roots in medicinal plant Coleus blumei

Document Type : Original Article


1 Department of Breeding and Biotechnology, Faculty of Agriculture, University of Tabriz, Tabriz, Iran

2 University of Farhangian, Tabriz, Iran



Hairy root induction in plants has affected by many factors including type, age of explant and bacterial strain. In the present study, the effects of bacterial strains (A4, C58, 15834 and GM), two concentrations (OD600= 0.5 and 1) and two explants (leaves and stems) in hairy root induction of C. blumei were investigated. According to the results of this study, the four strains showed different abilities to induce hairy root. The highest percent of induced roots (85%) was seen in explant treated with C58 strain, at OD600= 0.5 and the lowest induced roots (5%) by using of GM strain at OD600 = 1 and stem explants observed. The highest number of hairy roots per explant (12.94) has shown in leaf explant of leaf inoculated with C58 strain at (OD600 = 0.5) and the lowest number of hairy roots (1/1) was obtained from stem segments treated with GM strain at (OD600 = 1). According to the results of the analysis of variance, the effects of double reciprocal of strain in the concentration and strain in the explant was significant at a 1% probability level. Polymerase Chain Reaction (PCR) analysis using the role gene was performed for identification of the transformed hairy roots. In the lines of transgenic hairy roots, a sharp band of 304bp was amplified, but no such amplicon was observed in the untransformed root (negative control) sample. The results of this study indicated that the type of bacteria strain, explant type, and bacterial concentration are important factors in hairy roots induction.


Ayala-Silva, T., CA. Bey and G. Dortch. 2007. 'Agrobacterium rhizogenes mediated transformation of Asimina triloba L. cuttings'. Pakistan Journal of Biology Science, 10:132-13.
Bauer, N., D. Leljak-Levanić., S. Mihaljević and S. Jelaska. 2002. 'Genetic transformation of Coleus blumei Benth'. using Agrobacterium'. Food Technol. Biotechnol. 40: 163–169.
Bauer, N., D. Leljak-Levanić and S. Jelaska. 2004. 'Rosmarinic acid synthesis in transformed callus culture of Coleus blumei Benth'. Z. Naturforsch. 59:554–560.
Bertani, G. 1952. 'Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli'. J Bacteriol, 62:293-300.
Dellaporta, S., J. Wood and J.B. Hicks. 1983. 'A plant DNA mini preparation: Version II'. Plant Molecular Biological Reports, 1: 19-21.
Guillon, S., J. Trémouillaux-Guiller., PK. Pati., M. Rideau and P. Gantet. 2006a. 'Harnessing the potential of hairy roots: dawn of a new era'. Trends Biotechnol, 24: 403–409
Guillon, S., J. Trémouillaux-Guiller., PK. Pati., M. Rideau and P. Gantt. 2006b. 'Hairy root research: recent scenario and exciting prospects'. Curr Opin Plant Biol, 9: 341–346.
Hamilton, E. S., Cushnie, T. P. and Lamb, J. 2003. 'Assessment of the antibacterial activity of selected flavonoids and consideration of discrepancies between previous reports'. Microbiol. Res., 158: 281–289.
Herodez, S., S. Hadolin., M. Skerget and Z. Knez. 2003. 'Solvent extraction study of antioxidants from Lemon Balm (Melissa officinalis L.) Leaves'. Food Chem., 80: 275– 282.
Kumar, V., B. Jones and MR. Davey. 1991. 'Transformation by Agrobacterium rhizogenes and regeneration of transgenic shoots of the wild soybean Glycine argyrea'. Plant Cell Rep, 10:135-138.
Kuzovkina, I.N and B. Schneider. 2006. 'Progress in Botany'. Springer, Berlin, Heidelberg. 275-314.
Mary, AL 2005. 'Valuable secondary products from in vitro culture'. CRC Press LLC. 285-288
Murashige, T and F. Skoog. 1962. 'A revised medium for rapid growth and bioassays with tobacco tissue culture'. Physiol. Plant. 15, 473–497.
Namdeo, A. G., S. Patil and D. P. Fulzele. 2002. 'Influence of fungal elicitors on production of ajmalicine by cell cultures of Catharanthus roseus'. Biotechnol Prog, 18: 159-162.
Naderian, H., H. Ghahari and Sh. Asgari. 2012. 'Species diversity of natural enemies in corn fields and surrounding grasslands of Semnan province'. Iran. Calodema, 217, 1-8.
Nguyen, C., F. Bourgaud., P. Forlot and A. Guckert. 1992. 'Establishment of hairy root cultures of Psoralea species'. Plant Cell Reports, 11: 424-427.
Lee, S.Y., S.G. Kim, W.S. Song, Y.K. Kim and N.I. Park.2010. 'Influence of different strains of Agrobacterium rhizogenes on hairy root induction and production of alizarin and purpurin in Rubia akane'. Nakai. Rom. Biotechnol. Lett, 15: 5405-5409.
Palazon, J., A. Mallol., R. Eibl., C. Lettenbauer., RM. Cusido and MT. Pinol. 2003. 'Growth and ginsenoside production in hairy root cultures of Panax ginseng using a novel bioreactor'. Planta Med, 69: 344‑9.
Pirian, K., K. Piri and T. Ghiyasvand. 2012. 'Hairy roots induction from Portulaca oleracea using Agrobacterium rhizogenes to Noradrenaline’s production'. International Research Journal of Applied and Basic Sciences, 3(3): 642-649.
Samadi, A., J. Carapetian., R. Heidari., M. Jafri and A. Hassanzadeh. 2012.  'Hairy root induction in Linum mucronatum ssp. mucronatum, an anti-tumor lignans producing plant'. Not Bot Horti Agrobo, 40(1): 125-131.
Scheckel, K. A., S.C. Degner and D.F. Romagnolo. 2008. 'Rosmarinic acid antagonizes activator protein-1–dependent activation of cyclooxygenase-2 expression in human cancer and nonmalignant cell line'. J. Nutr, 138: 2098-2104.
Sevon, N and KM. OksmanCaldentey. 2002. 'Agrobacterium rhizogenes mediated transformation: Root cultures as a source of alkaloids'. Planta Med, 68:859‑68.
Shahabzadeh, Z., B. Heidari and R. Faramarzi Hafez. 2013. 'Induction of transgenic hairy roots in Trigonella foenumgraceum co-cultivated with Agrobacterium rhizogenes harboring a GFP Gene'. Journal of Crop Science Biotechnology, 16:263-268.
Shuai, L., Su. Liangchen, L. Shuai, Z. Xiaojun, Z. Danmin, L. Hong and L. Ling. 2016. 'Agrobacterium rhizogenes-mediated transformation of Arachis hypogaea: an efficient tool for functional study of genes.' Biotechnology & Biotechnological Equipment, 30:5, 869-878.
Sreeramanan, S., B. Vinod and S. Sashi. 2008. 'Optimization of the transient Gus a gene transfer of Phalaenopsis violacea orchid via Agrobacterium tumefaciens: an assessment of factors influencing the efficiency of gene transfer mechanisms'. Adv Nat Appl Sci, 2(2):77-88.
Sook Young L., H. Xu, Y. K. Kim and U.P. Sang, 2008. 'Rosmarinic acid production in hairy root cultures of Agastache rugosa Kuntze'. World J Microbiol Biotechnol, 24:969–972.
Vanishree, M., C. Lee., Y.S.F. Lo., S.M. Nalawade., C.Y. Lin and H.S. Tsay. 2004. 'Studies on the production of some important metabolites from medicinal plants by plant tissue cultures'. Bot Bull Acad Sin, 45: 1-22, (2004).
Vanhala, L., R. Hiltunen and K.M. Oksman-Caldentey. 1995. 'Virulence of different Agrobacterium strains on hairy root formation of Hyoscyamus muticus'. Plant Cell Reports, 14: 236-240.