The aim of this work was to construct transgenic plants with increased capabilities to degrade organic pollutants, such as polychlorinated biphenyls. The environmentally important gene of bacterial dioxygenase, the bphC gene, was chosen to clone into a plant of Nicotiana tabacum. The chosen bphC gene encodes 2,3-dihydroxybiphenyl-1,2-dioxygenase, which cleaves the aromatic ring of dihydroxybiphenyl, and we cloned it in fusion with the gene for β-glucuronidase (GUS), luciferase (LUC) or with a histidine tail. Several genetic constructs were designed and prepared and the possible expression of desired proteins in tobacco plants was studied by transient expression. We used genetic constructs successfully expressing dioxygenase's genes we used for preparation of transgenic tobacco plants by agrobacterial infection. The presence of transgenic DNA , mRNA and protein was determined in parental and the first filial generation of transgenic plants with the bphC gene. Properties of prepared transgenic plants will be further studied.

ICT Prague Faculty of Food and Biochemical Technology Department of Biochemistry and Microbiology Prague Czech Republic

Novakova M, Mackova M, Chrastilova Z, Viktorova J, Szekeres M, Demnerova K, et al. Cloning of the bacterial bphC gene into Nicotiana tabacum to improve the effciency of PCB phytoremediation. Biotechnol Bioeng. 2009;102:29–37. PubMed

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Golden R, Doull J, Waddell W, Mandel J. Potential human cancer risks from exposure to PCBs: a tale of two evaluations. Crit Rev Toxicol. 2003;33:543–580. PubMed

Macek T, Francova K, Sura M, Mackova M. Genetically modified plants with improved properties for phytoremediation purposes. In: Morel JL, Echevaria G, Goncharova N, editors. Phytoremediation of Metals. Dordrecht: NATO Science Series, Kluwer Academic Publishers; 2006. pp. 85–108.

Macek T, Uhlik O, Jecna K, Novakova M, Lovecka P, Rezek J, et al. Advances in phytoremediation and rhizoremediation. In: Singh A, editor. Advances in Applied Bioremediation. Springer-Verlag Berlin Heidelberg; 2009. pp. 257–277.

Schnoor JL, Licht LA, McCutcheon SC, Wolfe NL, Carreira LH. Phytoremediation of organic and nutrient contaminants. Environ Sci Technol. 1995;29:318–323. PubMed

Macek T, Mackova M, Kas J. Exploitation of plants for the removal of organics in environmental remediation. Biotechnol Adv. 2000;18:23–34. PubMed

Macková M, Barriault D, Francova K, Sylvestre M, Moder M, Vrchotová B, et al. Phytoremediation of polychlorinated biphenyls. In: Macková M, Dowling DN, Macek T, editors. Phytoremediation and Rhizoremediation. Springer-Verlag Berlin Heidelberg; 2006. pp. 143–167.

Macková M, Vrchotová B, Francová K, Sylvestre M, Tomaniová M, Lovecká P, et al. Biotransformation of PCBs by plants and bacteria—consequences of plant-microbe interactions. Eur J Soil Biol. 2007;43:233–241.

Leigh MB, Prouzova P, Mackova M, Macek T, Nagle DP, Fletcher JS. Polychlorinated biphenyls degrading bacteria associated with the trees in a PCB-contaminated site. Appl Environ Microbiol. 2006;72:2331–2342. PubMed PMC

Rezek J, der Wiesche C, Macková M, Zadražil F, Macek T. Biodegradation of PAHs in long-term contaminated soil cultivated with european white birch (Betula pendula) and red mulberry (Morus rubra) tree. Int J Phytorem. 2009;11:1–16.

Demnerova K, Mackova M, Spevakova V, Beranova K, Kochankova L, Lovecka P, et al. Two approaches to biological decontamination of groundwater and soil polluted by aromatics-characterization of microbial populations. Int Microbiol. 2005;8:205–211. PubMed

Ionescu M, Beranova K, Dudkova V, Kochankova L, Demnerova K, Macek T, et al. Isolation and characterization of different plant associated bacteria and their potential to degrade polychlorinated biphenyls. Int Biodeterior Biodegrad. 2009;63:667–672.

Mackova M, Prouzova P, Stursa P, Ryslava E, Uhlik O, Beranova K, et al. Phyto/rhizoremediation studies using long-term PCB-contaminated soil. Environ Sci Pollut Res. 2009;16:817–829. PubMed

Uhlik O, Jecna K, Leigh MB, Mackova M, Macek T. DNA-based stable isotope probing: a link between community structure and function. Sci Total Environ. 2009;407:3611–3619. PubMed

Uhlik O, Jecna K, Mackova M, Vlcek C, Hroudova M, Demnerova K, et al. Biphenyl-metabolizing bacteria in the rhizosphere of horseradish and bulk soil contaminated by polychlorinated biphenyls as revealed by stable isotope probing. Appl Environ Microbiol. 2009;75:6471–6477. PubMed PMC

Novakova M, Mackova M, Chrastilova Z, Viktorova J, Szekeres M, Demnerova K, et al. Cloning of the bacterial bphC gene into Nicotiana tabacum to improve the efficiency of PCB phytoremediation. Biotechnol Bioeng. 2009;102:29–37. PubMed

Fletcher JS, Donnelly PK, Hedge RS. Biostimulation of PCB-degrading bacteria by compounds released from plant roots. In: Hinchee RE, Anderson DB, Hoeppel RE, editors. Bioremediation of Recalcitrant Organics. Columbus: Battelle Press; 1995. pp. 131–136.

Kucerova P, Mackova M, Chroma L, Burkhard J, Triska J, Demnerova K, et al. Metabolism of polychlorinated biphenyls by Solanum nigrum hairy root clone SNC-90 and analysis of transformation products. Plant Soil. 2000;225:109–115.

Wilken A, Bock C, Bokern M, Harms H. Metabolism of different PCB congeners in plant cell cultures. Environ Chem Toxicol. 1995;14:2017–2022.

Rezek J, Macek T, Mackova M, Triska J. Plant metabolites of polychlorinated biphenyls in hairy root culture of black nightshade Solanum nigrum SNC-90. Chemosphere. 2007;69:1221–1227. PubMed

Lovecká P, Melenová I, Kucerová P, Nováková H, Macková M, Ruml T, et al. Products of biological PCB degradation and their ecotoxicity. Int Biodeterior Biodegrad. 2004;53:195–283.

Lovecká P, Macková M, Demnerová K. Metabolic products of PCBs in bacteria and plants-comparison of their toxicity and genotoxicity. In: Fava F, Canepa P, editors. Innovative approaches to the bioremediation of contaminated sites. Venice: INCA; 2005. p. 407.

Seeger M, Timmis KN, Hofer B. Conversion of chlorobiphenyls into phenylhexadienoates and benzoates by the enzymes of the upper pathway for polychlorobiphenyl degradation encoded by the bph locus of Pseudomonas sp strain LB400. Appl Environ Microbiol. 1995;61:2654–2658. PubMed PMC

Abramowicz DA. Aerobic and anaerobic biodegradation of PCBs: a review. Crit Rev Biotechnol. 1990;10:241–251.

Furukawa K. Molecular genetics and evolutionary relationship of PCB-degrading bacteria. Biodegradation. 2004;5:289–300. PubMed

Mondello FJ, Turcich MP, Lobos JH, Erickson BD. Identification and modification of biphenyl dioxygenase sequence that determine the specificity of polychlorinated biphenyl degradation. Appl Environ Microbiol. 1997;63:3096–3103. PubMed PMC

Sondossi M, Barriault D, Sylvestre M. Metabolism of 2,2′- and 3,3′-dihydroxybiphenyl by the biphenyl catabolic pathway of Comamonas testosteroni B-356. Appl Environ Microbiol. 2004;70:174–181. PubMed PMC

Mohammadi M, Chalavi V, Novakova-Sura M, Laliberte JF, Sylvestre M. Expression of bacterial biphenyl-chlorobiphenyl dioxygenase genes in tobacco plants. Biotechnol Bioeng. 2007;97:496–505. PubMed

Erickson BD, Mondello FJ. Nucleotide sequencing and transcriptional mapping of the genes encoding biphenyl dioxygenase, a multicomponent polychlorinated-biphenyl-degrading enzyme in Pseudomonas strain LB400. J Bacteriol. 1992;174:2903–2912. PubMed PMC

Taira K, Hirose J, Hayashida S, Furukawa K. Analysis of bph operon from the polychlorinated biphenyldegrading strain of Pseudomonas pseudoalcaligenes KF707. J Biol Chem. 1992;267:4844–4853. PubMed

Sylvestre M, Sirois M, Hurtubise Y, Bergeron J, Ahmad D, Shareck F, et al. Sequencing of Comamonas testosteroni strain B-356-biphenyl/chlorobiphenyl dioxygenase genes: evolutionary relationships among Gram-negative bacterial biphenyl dioxygenases. Gene. 1996;174:195–202. PubMed

Rezek J, Macek T, Macková M, Triska J, Ruzickova K. Hydroxy-PCBs, methoxy-PCBs and hydroxymethoxy-PCBs: Metabolites of polychlorinated biphenyls formed in vitro by tobacco cells. Environ Sci Technol. 2008;42:5746–5751. PubMed

Macek T, Kotrba P, Svatos A, Novakova M, Demnerova K, Mackova M. Novel roles for genetically modified plants in environmental protection. Trends Biotechnol. 2008;26:146–152. PubMed

Sandermann H. Higher plant metabolism of xenobiotics: the “green liver” concept. Pharmacogenetics. 1994;4:225–241. PubMed

Cherian S, Oliveira MM. Transgenic plants in phytoremediation: Recent advances and new possibilities. Environ Sci Technol. 2005;39:9377–9390. PubMed

Francová K, Macková M, Macek T, Sylvestre M. Ability of bacterial biphenyl dioxygenases from Burkholderia sp. LB400 and Comamonas testosteroni B-356 to catalyse oxygenation of ortho-hydroxychlorobiphenyls formed from PCBs by plants. Environ Pollut. 2004;127:41–48. PubMed

Sylvestre M. Genetically modified organisms to remediate polychlorinated biphenyls. Where do we stand? Int Biodeterior Biodegrad. 2004;54:153–162.

Uchida E, Ouchi T, Suzuki Y, Yoshida T, Habe H, Yamaguchi I, et al. Secretion of bacterial xenobiotic-degrading enzymes from transgenic plants by an apoplastic expressional system: an applicability for phytoremediation. Environ Sci Technol. 2005;39:7671–7677. PubMed

Hein P, Powlowski J, Barriault D, Hurtubise Y, Ahmad D, Sylvestre M. Biphenyl-associated metacleavage dioxygenases from Comamonas testosteroni B-356. Can J Microbiol. 1998;44:42–49. PubMed

Borovka R, Szekeres M, Macek T, Kotrba P, Sylvestre M, Mackova M. First steps in attempt to enhance the ability of plants to metabolise polychlorinated biphenyls by introduction of bacterial genes to plant DNA. Int Biodeterior Biodegrad. 1998;42:243.

Surá M, Macková M, Borovka R, Francová K, Szekeres M, Macek T. Cloning of bacterial PCBdegrading gene into the plants. In: Verstraete W, editor. Environmental Biotechnology. ESEB 2004. London: Taylor and Francis Group; 2004. pp. 749–752.

Sylvestre M, Macek T, Mackova M. Transgenic plants to improve rhizoremediation of polychlorinated biphenyls (PCBs) Curr Opin Biotechnol. 2009;20:242–247. PubMed

Macek T, Surá M, Pavlíková D, Francová K, Scouten WH, Szekeres M, et al. Can tobacco have potentially beneficial effect to our health? Z Naturforsch C. 2005;60:292–299. PubMed

Kotrba P, Najmanova J, Macek T, Ruml T, Mackova M. Genetically modified plants in phytoremediation of heavy metal and metalloid soil and sediment pollution. Biotechnol Adv. 2009;27:799–810. PubMed

Hannink R, Rosser SJ, French ChE, Basran A, Murray JAH, Nicklin S, et al. Phytodetoxification of TNT by transgenic plants expressing a bacterial nitroreductase. Nat Biotechnol. 2001;19:1120–1121. PubMed

VanAken B, Correa P, Schnoor J. Phytoremediation of polychlorinated biphenyls: new trends and promises. Environ Sci Technol. 2010;44:2767–2776. PubMed PMC

VanAken B. Transgenic plants for phytoremediation: Helping nature to clean up environmental pollution. Trends Biotechnol. 2008;26:225–227. PubMed