Optimization of hairy root culture establishment in Chicory plants (Cichorium intybus) through inoculation by Agrobacterium rhizogenes

Document Type : Research Paper

Authors

Abstract

Agrobacterium rhizogenes induced hairy roots gain of the potential of production and accumulation of plant secondary metabolites. In recent decades, many researchers have focused on the biosynthesis of valuable secondary metabolites in hairy roots, because genetically modified roots produce such compounds with more genetic and biosynthetic stability than cell suspension cultures. Chicory (Cichorium intybus L.) is a biennial or perennial medicinal plant from Asteraceae. The plant contains many important metabolites including inulin, scoline, coumarin, vitamins, flavonoids and aromatics. In this study, with the aim of optimizing hairy root culture establishment, cotyledons of Chicory plants inoculated with different strains of Agrobacterium rhizogenes were cultured on different plant tissue culture media. Numbers of hairy roots produced in different experiments were recorded and molecular confirmation of hairy root clones was performed through specific amplification of rolB and rolC genes in PCR reactions. Based on the efficiency of hairy root induction in different experiments, the best combination of bacterial strain-culture medium for establishing hairy root cultures of Chicory were determined. Agrobacterium rhizogenes strains A4, A13 and 15834 induced hairy roots in Chicory and MS medium was considered as the best culture medium for developing and establishment of hairy root cultures.
 

Keywords


Agrobacterium rhizogenes, hairy root, Chicory, secondary metabolites

Azlan GJ, Marziah M, Radzali M, Johari R (2002). Establishment of Physalis minima hairy roots culture for the production of Physalins. Plant Cell, Tissue and Organ Culture 69: 271-278. 
Barik DP, Mohapatra U, Chand PK (2005). Transgenic grasspea (Lathyrus sativus  L.): factors influencing  Agrobacterium-mediated transformation and regeneration.   Plant Cell Reports 24: 523-531.
Baron C, Domke N, Beinhofer M, Hapfelmeier S (2001). Elevated temperature differentially affects virulence, VirB protein accumulation, and T-pilus formation in different  Agrobacterium tumefaciens and Agrobacterium vitis strains.  Journal of Bacteriology 183: 6852-6861.
Cao D, Hou W, Song S, Sun H, Wu C, Gao Y, Han T (2009). Assessment of conditions affecting Agrobacterium rhizogenes-mediated transformation of soybean. Plant Cell Tissue Organ Cult 96: 45-52.
Chabaud M, de Carvalho-Niebel F, Barker DG (2003). Efficient transformation of Medicago truncatula cv. Jemalong using the hyper virulent Agrobacterium tumefaciens  strain AGL1. Plant Cell Reports  22: 46-51.
Chen L, Zhang B, Z Xu (2008). Salt tolerance conferred byoverexpression of  Arabidopsis  vacuolar Na(+)/H(+) antiporter geneAtNHX1 in common buckwheat (Fagopyrum esculentum).Transgenic Research 17: 121-132.
Condori J, Sivakumar G, Hubstenberger J, Dolan M, Sobolev V, Medina-Bolivar F (2010). Induced biosynthesis of resveratrol and the prenylated stilbenoidsarachidin-1 and arachidin-3 in hairy root cultures of peanut: effects of culture medium and growth stage.  Plant Physiol. Biochem 48: 310–318.
Corbin DR, Grebenok RJ, Ohnmeiss TE, Green PJT Purcell JP (2001). Expression and chloroplast targeting of cholesterol oxidase in transgenic tabacco plants. Plant physiology 126: 1116-1128.
Crane C, Wright E, Dixon RA, Wang ZY (2006).  Transgenic Medicago truncatula plants obtained from Agrobacterium tumefaciens-transformed roots and  Agrobacterium rhizogenes-transformed hairy roots.  Planta 223: 1344-1354.
De Buck S, Jacobs A, Van Montagu M, Depicker A (1998). Agrobacterium tumefaciens  transformation and cotransformation frequencies of Arabidopsis thaliana root explants and tobacco protoplasts.  Mol Plant Microbe Interact 11: 449-457.
Doran PM (2002). Properties and applications of hairy-root cultures. Plant Biotechnology  and Transgenic Plants (Eds. K. M. okasman-caldenty and W. H. Barz). Mercel dekker Inc.new York. 4: 143-162
Geier T, Sangwan RS (1996).  Histology and chimeral segregationreveal cell-specific differences in the competence for shoot regeneration and Agrobacterium-mediated transformation in Kohleria internode explants.  Plant Cell Reports 15: 386-390.
Giri A, Narasu ML (2000). Transgenic hairy root: recent trends and application Biotechnology      advances 18: 1-22.
Kim KH, Lee YH, Kim D, Park YH, Lee JY, Hwang YS, Kim YH (2004). Agrobacterium-mediated genetic transformation of Perilla frutescens.  Plant Cell Reports 23: 386-390.
Krolicka A, Stanszewska I, Bielawski K, Malinski E, Szafranek J, Lojkowska E  (2001). Establishment of hairy roots of Ammimajus. Plant Science 160: 259-264.
Kumar V, Sharma A, Prasal B, Chayapathy N, Gururaj HB, Ravishahnkar GA (2006). A.rhizogenes mediated genetic transformation resulting in hairy root formation is enhanced by ultrasonication and acetosyringone treatment.  Electronic Journal of Biotechnology 9: 143-151.
Lee MH, Yoon ES, Jeong JH, Choi YE (2004). Agrobacterium rhizogenes-mediated transformation of Taraxacum platycarpum and changes of morphological characters.  Plant Cell Reports 22: 822-827.
Mano Y, Ohkawa H, Yamada Y (1989). Production of tropane alkaloid by hairy root cultures of Duboisia leichhardtii transformed by Agrobacterium rhizogenes. Plant Science 59: 191–201.
Mao  Q,  Sultan  H,  Tursun  S,  Mavlanja  (2009).  Determination  of  Total Flavonoids from Cichorium intybus L. Biotechnology 19: 78-80.
Munoz  CLM 2004.  Spanish  medicinal  Plants:  Cichorium  intybus  L. Boletin de la RealSociedad Espanola de Historia Natural 99: 41-47.
Otani M, Mii M, Handa T, Kamada H, Shimada T (1993). Transformation of sweet potato(Ipomoea batatas (L.) Lam.) plants by Agrobacterium rhizogenes. Plant Sci 94: 151-159.
Saito K, Yamazaki M, Anzai H, Yoneyama K, Murakoshi I (1992). Transgenic herbicide-resistant Atropa belladonna using an Ri binary vector and inheritance of the transgenic trait. Plant Cell Rep 11: 219-224.
Sevon N, Hiltunen R, Oksman Caldentey KM (1998). Somaclonal variation in Agrobacterium transformed roots and protoplast-derived hairy root clones of Hyoscyamusmuticus. Planta Med. 64: 37-41.
Shakti M, Arun KK, Suman PSK, Bhartendu NM (2008). Genetic transformation studies and scale up of hairy root culture of Glycyrrhiza glabra in bioreactor.  Electronic Journal of Biotechnology 11: 0717-3458.
Shen WH, Petit A, Guern, J,  Tempe J (1988). Hairy roots are more sensitive to auxin than normal roots. Proceeding of the National Academy of Sciences 85: 3417-3421.
Shinde A, Malpathak N, Fulzele D (2010). Impact of nutrient components onproduction of the phytoestrogens daidzein and genistein by hairy roots of Psoralea corylifolia. journal of natural medicines 64: 346–353.
Shrutika Dhakulkar TR, Ganapathi SB (2005). Induction of hairy roots in Gmelina arborea Roxb. and production of verbascoside in hairy roots. Plant Science 169: 812–818
Tamakawa T, Sekiguchi S, Kodama T, Smith S, Yeoman MM (1998). Transformation of Chilli Pepper (Capsicum frutescens) With a Phenylalanine Amnlonia-Lyase Gene.Plant Biotechnolog 15: 189-193.
Tepfer D (1984). Transformation of several species of highplants by Agrobacterium rhizogenes: Sexual transmission of the transformed genotype and phenot Cell 37: 959- 967.
Tomilov A, Tomilova N, Yoder JI (2007). Agrobacterium tumefaciens and Agrobacterium rhizogenes transformed roots of the parasitic-plant  Triphysaria versicolor  retain parasitic competence .Planta  225: 1059-1071.
Vanhala L, Hiltunen R, Oksman-Caldentey KM (1995). Virulence of different  agrobacterium strains  on hairy root formation of Hyoscyamus muticus. Plant cell Rep 14: 236-240.
Visser RGF, Jacobsen E, Witholt B, Feenstra WJ (1989). Efficient transformation of potato(Solanum tuberosum L.) using binary vector in Agrobacterium rhizogenes. Theor. Appl. Genet 78: 594-600.
Woodhead  M,  Davies  HV,  Brennan  RM, Taylor MA  (1998). The isolation of genomic DNA  from black currant  (Ribes nigrum L.). Mol. Biotechnol 9: 243-246.
Yamazaki M, Son L, Hayashi T, Morita N, Asamizu T, Murakoshi I, Saito K (1996). Transgenic fertile Scoparia dulcis L., a folk medicinal plant, conferred with a herbicide-resistant trait using an Ri binary vector. Plant Cell Rep 15: 317-321.
Yang Y (2009). Process  Optimization  of  Extracting  Phenols  from Cichorium  intybus cv. Puna with Response Surface Methodology. J. Northwest Forest  University 24:  118-120.