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Effects of catechins on Agrobacterium-mediated genetic transformation of Camellia sinensis

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Abstract

In this study, recalcitrance of tea plant ( Camellia sinensis) to Agrobacterium-mediated genetic transformation was investigated with an emphasis on specialized compounds in tea. Chemical constituents in tea leaves and calli were extracted into liquid Luria–Bertani (LB) medium to determine their biological activities on Agrobacterium growth, virulence, and plant transformation efficiency. Compared to the control Agrobacterium grown in LB medium, tea leaf extract containing 6.5 mg mL−1 catechins resulted in an 84.6 % reduction of Agrobacterium growth, a 73–36 % suppression of expression for the six virulence (vir) genes, browning of infected tobacco explant wounds, and an absence of transient or stable transformation events. Tea callus extract, containing 0.22 mg mL−1 catechins, did not significantly affect Agrobacterium growth or tobacco transgenic hairy root generation, whereas it enhanced the expression of some vir genes. Treatment with authentic catechin mixtures (other than caffeine) dissolved in LB resulted in suppression of Agrobacterium growth, vir gene expression, and tobacco transformation efficiency. Our data suggest that catechins are the key active constituents in tea leaves. Transient transformation efficiencies of tea leaves were much lower than those of tobacco leaves as indicated by the GUS (β-glucuronidase) assay, probably a result of inhibition by the catechins present in tea leaves. Lower transformation efficiencies of tea calli suggested that additional plant factor(s) might also exert inhibitory effects on tea plant transformation. Agrobacterium rhizogenes ATCC 15834 induced transgenic roots from the tea explants with 15–20 % efficiency. Our data suggested catechins inhibition of tea gene transformation could be overcome by using optimized strains of Agrobacterium.

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Abbreviations

HPLC:

High performance liquid chromatography

AS:

Acetosyringone

GUS:

β-glucuronidase

EDTA:

Ethylene diamine tetraacetic acid

PCR:

Polymerase chain reaction

References

  • Ahn YJ, Kawamura T, Kim M, Yamamoto T, Mitsuoka T (1991) Tea polyphenols: selective growth inhibitors of Clostridium spp. Agric Biol Chem 55:1425–1426

    Article  CAS  Google Scholar 

  • Amita B, Priyanka S, Vitaly C (2010) The roles of plant phenolics in defence and communication during Agrobacterium and Rhizobium infection. Mol Plant Pathol 11:705–719

    Google Scholar 

  • Anand A, Krichevsky A, Schornack S, Lahaye T, Tzfira T, Tang Y, Citovsky V, Mysore KS (2007) Arabidopsis VirE2 interacting protein2 is required for Agrobacterium T-DNA integration in plants. Plant Cell 19:1695–1708

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Ashby AM, Watson MD, Loake GJ, Shaw CH (1988) Tiplasmid specific chemotaxis of Agrobacterium tumefaciens C58C1 towards vir inducing phenolic compounds and soluble factors from mono cotyledons and dicotyledonous plants. J Bacteriol 170:4181–4187

    PubMed  CAS  PubMed Central  Google Scholar 

  • Balentine DA, Wiseman SA, Bouwens LCM (1997) The chemistry of tea flavonoids. Crit Rev Food Sci 37:693–704

    Article  CAS  Google Scholar 

  • Collin HA, Edwards S (1998) Plant cell culture. BIOS Scientific Publisher Limited 1:10–15

  • Dada KD, Tripathy L (2005) Agrobacterium-induced hypersensitive necrotic reaction in plant cells: a resistance response against Agrobacterium-mediated DNA transfer. Afr J Biotechnol 4:752–757

    Google Scholar 

  • Deng WW, Wang S, Chen Q, Zhang ZZ, Hu XY (2012) Effect of salt treatment on theanine biosynthesis in Camellia sinensis seedlings. Plant Physiol Biochem 56:35–40

    Article  PubMed  CAS  Google Scholar 

  • Dixon RA, Pasinetti GM (2010) Flavonoids and isoflavonoids: from plant biology to agriculture and neuroscience. Plant Physiol 154:453–457

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Dunnett CW (1955) A multiple comparison procedure for comparing several treatments with a control. J Am Statist Assoc 50:1096–1121

    Article  Google Scholar 

  • Feng L, Hou RY, Zhang L, Wan XC, Gao MJ, Wei S (2014) Determination of quality constituents in the young leaves of albino tea cultivars. Food Chem 155:98–104

    Article  PubMed  CAS  Google Scholar 

  • Forrest GI (1969) Studies on the polyphenol metabolism of tissue cultures derived from the tea plant [Camellia sinensis (L.)]. Biochem J 113:765–772

    PubMed  CAS  PubMed Central  Google Scholar 

  • Fronzes R, Christie PJ, Waksman G (2009) The structural biology of type IV secretion systems. Nat Rev Microbiol 7:703–714

    Article  PubMed  CAS  Google Scholar 

  • Gamborg OL, Miller RA, Ojima K (1968) Nutrient requirements of suspension culture of soybean root cells. Exp Cell Res 50:151–158

    Article  PubMed  CAS  Google Scholar 

  • Gelvin SB (2000) Agrobacterium and plant genes involved in T-DNA transfer and integration. Annu Rev Plant Physiol Plant Mol Biol 51:223–256

    Article  PubMed  CAS  Google Scholar 

  • Hooykaas PJJ, Klapwijk PM, Nuti MP, Schilperoort RA, Rörsch A (1977) Transfer of the Agrobacterium tumefaciens TI plasmid to avirulent Agrobacteria and to Rhizobium ex planta. J Gen Microbiol 98:477–484

    Article  Google Scholar 

  • Horsch RB, Fry JE, Hoffman NL, Eichholtz D, Rogers SG, Fraley RT (1985) A simple and general method for transferring genes into plants. Science 227:1229–1231

    Article  CAS  Google Scholar 

  • Janssen BJ, Gardner RC (1989) Localized transient expression of GUS in leaf discs following cocultivation with Agrobacterium. Plant Mol Biol 14:61–72

    Article  Google Scholar 

  • John KMM, Joshi SD, Mandal AKA, Kumar SR, Kumar RR (2009) Agrobacterium rhizogenes-mediated hairy root production in tea leaves [Camellia sinensis (L.) O. Kuntze]. Indian J Biotechnol 8:430–434

    CAS  Google Scholar 

  • Kanwar J, Taskeen M, Mohammad I, Huo C, Chan TH, Dou QP (2012) Recent advances on tea polyphenols. Front Biosci (Elite Ed) 4:111–131

    Article  Google Scholar 

  • Kumar N, Pandey S, Bhattacharya A, Ahuja PS (2004) Do leaf surface characteristics affect Agrobacterium infection in tea (Camellia sinensis (L.) O. Kuntze). J Bioscience 29:309–317

    Article  Google Scholar 

  • Li XB, Cai L, Cheng NH, Liu JW (2002) Molecular characterization of the cotton GhTUB1 gene that is preferentially expressed in fiber. Plant Physiol 130:666–674

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Liu YJ, Gao LP, Xia T, Zhao L (2009) Investigation of the site-specific accumulation of catechins in the tea plant (Camellia sinensis (L.) O. Kuntze) via vanillin-HCl staining. J Agric Food Chem 57:10371–10376

    Article  PubMed  CAS  Google Scholar 

  • Livak K, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCt method. Methods 25:402–408

    Article  PubMed  CAS  Google Scholar 

  • Lopez SJ, Kumar RR, Pius PK, Muraleedharan N (2004) Agrobacterium tumefaciens-mediated genetic transformation in tea (Camellia sinensis [L.] O. Kuntze). Plant Mol Biol Rep 22:201–202

    Article  Google Scholar 

  • Luo YY, Liang YR (2000) Studies on the construction of Bt gene expression vector and its transformation in tea plant. J Tea Sci 20:141–147

    CAS  Google Scholar 

  • Matsumoto S, Fukui M (1998) Agrobacterium tumefaciens medaited gene transfer in tea plant (Camellia sinensis) cells. Jpn Agric Res Q 32:287–291

    CAS  Google Scholar 

  • Medina-Bolivar F, Condori J, Rimando AM, Hubstenberger J, Shelton K, O’Keefe SF, Bennett S, Dolan MC (2007) Production and secretion of resveratrol in hairy root cultures of peanut. Phytochemistry 68:1992–2003

    Article  PubMed  CAS  Google Scholar 

  • Mohanpuria P, Kumar V, Ahuja PS, Yadav SK (2011) Agrobacterium-mediated silencing of caffeine synthesis through root transformation in Camellia sinensis L. Mol Biotechnol 48:235–243

    Article  PubMed  CAS  Google Scholar 

  • Mohanpuria P, Rana NK, Yadav SK (2008) Transient RNAi based gene silencing of glutathione synthetase reduces glutathione content in Camellia sinensis (L.) O. Kuntze somatic embryos. Biol Plantarum 52(2):381–384

    Article  CAS  Google Scholar 

  • Mondal TK, Bhattacharya A, Ahuja PS, Chand PK (2001a) Transgenic tea [Camellia sinensis (L.) O. Kuntze cv. Kangra Jat] plants obtained by Agrobacterium-mediated transformation of somatic embryos. Plant Cell Rep 20:712–720

    Article  CAS  Google Scholar 

  • Mondal TK, Bhattacharya A, Ahuja PS, Chand PK (2001b) Factor effecting Agrobacterium tumefaciens mediated transformation of tea (Camellia sinensis (L). O.Kuntze). Plant Cell Rep 20:712–720

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Pang Y, Abeysinghe ISB, He J, He X, Huhman D, Mewan KM, Sumner LW, Yun J, Dixon RA (2013) Functional characterization of proanthocyanidin pathway enzymes from tea and their application for metabolic engineering. Plant Physiol 161:1103–1116

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Pitzschke A (2013) Agrobacterium infection and plant defense-transformation success hangs by a thread. Front Plant Sci 4:519

    Article  PubMed  PubMed Central  Google Scholar 

  • Sandal I, Kumar A, Bhattacharya A, Desikalhar RS, Gulati A, Ahuja PS (2001) A thermolabile caffeine fraction of tea leaves–A substitute of acetosyringone for Agrobacterium-mediated genetic transformations (Patent filed in US and PCT)

  • Sandal I, Saini U, Lacroix B, Bhattacharya A, Ahuja PS (2007) Agrobacterium-mediated genetic transformation of tea leaf explants: effects of counteracting bactericidity of leaf polyphenols without loss of bacterial virulence. Plant Cell Rep 26:169–176

    Article  PubMed  CAS  Google Scholar 

  • Schrammeijer B, Beijersbergen A, Idler KB, Melchers LS, Thompson DV, Hooykaas PJ (2000) Sequence analysis of the vir-region from Agrobacterium tumefaciens octopine Ti plasmid pTi15955. J Exp Bot 51:1167–1169

    Article  PubMed  CAS  Google Scholar 

  • Shibasaki-Kitakawa N, Takeishi J, Yonemoto T (2003) Improvement of catechin productivity in suspension cultures of tea callus cells. Biotechnol Prog 19:655–658

    Article  PubMed  CAS  Google Scholar 

  • Wu XX, Li J, Wang ZK, Liu SS, Li HH, Ma Y, Li WB (2010) Effect of AS concentration and pH on soybean genetic transformation. J Northeast Agric Univ 41:1–4

    Google Scholar 

  • Yam TS, Shah S, Hamilton-Miller JM (1997) Microbiological activity of whole and fractionated crude extract of tea (Camellia sinensis), and of tea components. FEMS Microbiol Lett 152:169–174

    Article  PubMed  CAS  Google Scholar 

  • Yuan ZC, Edlind MP, Liu P, Saenkham P, Banta LM, Wise AA, Ronzone E, Binns AN, Kerr K, Nester EW (2007) The plant signal salicylic acid shuts down expression of the vir regulon and activates quormone-quenching genes in Agrobacterium. Proc Natl Acad Sci USA 104:11790–11795

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Zupan JR, Zambryski P (1995) Transfer of T-DNA from Agrobacterium to the plant cell. Plant Physiol 107:1041–1047

    Article  PubMed  CAS  PubMed Central  Google Scholar 

Download references

Acknowledgments

We thank Prof Li-Ping Gao at the Anhui Agricultural University and Prof. Jianliang Lu at the Zhejiang University for providing tea calli and the Agrobacterium strains. This work was financially supported by the National Science Foundation in China (#31070614 and #31370687), the Doctoral Programs of Higher Education of the Ministry of Education (#20123418110002), the Program for Changjiang Scholars and Innovative Research Team in Universities (IRT1101), and the “Twelfth Five-Year” National Key Basic Research and Development Project (973) in China (2012CB722903).

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Correspondence to Shu Wei.

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Song, DP., Feng, L., Rana, M.M. et al. Effects of catechins on Agrobacterium-mediated genetic transformation of Camellia sinensis . Plant Cell Tiss Organ Cult 119, 27–37 (2014). https://doi.org/10.1007/s11240-014-0511-7

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  • DOI: https://doi.org/10.1007/s11240-014-0511-7

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