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Applications of Biotechnology: Phytoremediation

In this section we will look at how the tools of biotechnology can be applied to a specific problem. Phytoremediation: use of plants to remediate polluted soil and/or water. It is an alternative to engineering or processing on site. Applications of Biotechnology: Phytoremediation.

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Applications of Biotechnology: Phytoremediation

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  1. In this section we will look at how the tools of biotechnology can be applied to a specific problem. Phytoremediation: use of plants to remediate polluted soil and/or water. It is an alternative to engineering or processing on site. Applications of Biotechnology: Phytoremediation COURSE FIGURES CAN BE DOWNLOADED THROUGH http://www.courseweb.uottawa.ca/BIO4174

  2. BIOTECHNOLOGY: Phytoremediation • Outline: • Use of Plants for Bioremediation • Biodegradation of explosives • Biodegradation of organomercury • Bioaccumulation of arsenic • Future of Phytoremediation

  3. BIOTECHNOLOGY: Phytoremediation • Definitions: • Phytovolatilization: take up contaminants through the roots, • transport them to the leaves, release as a detoxified vapor to atmosphere. • Microorganism stimulation:root exudates stimulate growth of • microorganisms(fungi,bacteria) that metabolize the organic contaminants. • Phytostabilization:plants prevent contaminants from migrating by • reducing runoff, surface erosion, and ground-water flow rates. • Phytoaccumulation/extraction:roots remove metals from soil, • transport them to leaves & stems for harvesting or metal recovery. • Phytodegradation by plants:roots remove organics that are • metabolized to non-toxic molecules within the plant.

  4. BIOTECHNOLOGY: Phytoremediation From Pilon-Smits (2005) Annu Rev Plant Biol 56: 15-39

  5. BIOTECHNOLOGY: Phytoremediation Use of Plants for Bioremediation • Present strategies: • Bioaccumulation (transport and storage for harvest) • Bioprocessing (chemical transformation to CO2, NH3, Cl- & SO42-) • Advantages of plants: • Plants demonstrate tolerance to toxins • Photosynthesis-free energy • Extensive root systems to mine soil (440 million km/h/yr**) • Plants have evolved for transport of materials • Large biomass for harvesting • Species adapted to different ecosystems including wetlands • Disadvantages of plants: • Construction of transgenics • Requires release, opposition to GMOs

  6. BIOTECHNOLOGY: Phytoremediation Quotation from R. Meagher’s Lab (http://www.genetics.uga.edu/rbmlab/phyto/phyto.html ) “Phytoremediation is affordable on the grand scale needed for marginal land reclamation and cleaning the water in lakes, streams, and marshes.” • We will use three examples to illustrate phytoremediation: • Biodegradation of Explosives • Biodegredation of Organomercury • Bioaccumulation of Arsenic

  7. BIOTECHNOLOGY: Phytoremediation Biodegradation of explosives The Problem: Contamination to explosives TNT, RDX and glycerol trinitrate. “Exposure to TNT and RDX, and their degradation products causes symptoms such as anemia and liver damage. These chemicals can be lethal and are suspected carcinogens. Hundreds of tons of these compounds are found in sediments at innumerable manufacturing sites and storage sites for unexploded ordnance around the world. Tens of thousands of acres of land and water resources are unsafe because of RDX and TNT contamination.” The “Solution”: Engineer plants that are able to degrade these compounds in situ.

  8. BIOTECHNOLOGY: Phytoremediation Biodegradation of explosives These are the targets Breakdown of TNT to ADNT (monoaminodinitrotoluene) can create “sterile” pink water lagoons.

  9. BIOTECHNOLOGY: Phytoremediation • Biodegradation of explosives: • The general strategy is to isolate catabolic genes and through • standard cloning technologies convert them into plant genes expressed • via a • constitutive promoter such as the CaMV 35S promoter for expression • throughout the plant. • tissue-specific promoter for expression to leaves, stems etc. • Following transformation and plant regeneration, the properties of • the plant are evaluated by standard tests. • Bacteria are the usual source of the genes, bacteria such as • Enterobacter cloacae or Rhodococcus rhodochrous that can grow on these • contaminants.

  10. BIOTECHNOLOGY: Phytoremediation Biodegradation of explosives: Summary from Meagher (2006). Key: RDX, TNT, ADNT: See Fig 6 NR: nitroreductase XplA: RDX-degrading cytochrome P450 NDAB: 4-nitro-2,4-diazabutanal.

  11. BIOTECHNOLOGY: Phytoremediation Biodegradation of explosives: from Rylott et al. (2006) • Conclusion from these experiments: • decontamination works in a model system • TNT can be catabolized • RDX is catabolized and the nitrogen used for growth (win-win!!)

  12. BIOTECHNOLOGY: Phytoremediation Biodegredation of Organomercury Methylmercury is a pollutant that biomagnifies in the aquatic food chain with severe consequences for humans and other animals. The main targets include free cysteine in proteins and peptides leading to damage in the central nervous system. Symptoms include sensory impairment (vision, hearing, speech), disturbed sensation and a lack of coordination. Mercury occurs in deposits throughout the world and it is harmless in an insoluble form, such as mercuric sulfide, but it is poisonous as methylmercury [CH3Hg]+ due to its aqueous solubility. Sources of Mercury include burning coal and mineral extraction. Many uses of mercury are being curtailed or eliminated.

  13. BIOTECHNOLOGY: Phytoremediation Biodegredation of Organomercury: protein targeting

  14. BIOTECHNOLOGY: Phytoremediation Biodegredation of Organomercury: chloroplast targeting in tobacco. Key: WT = cv Petit Havana 5A = pLDR-MerAB 9 = pLDR-MerAB-3’-UTR

  15. 25 54 61 Volatilization assay with 5μM phenylmercury acetate. Samples are lines 22, 25, 54 & 61 BIOTECHNOLOGY: Phytoremediation Biodegredation of Organomercury by Transgenic Poplar data from Lyrra et al. (2007)

  16. BIOTECHNOLOGY: Phytoremediation Biodegredation of Organomercury: from Omichinski, 2007 Should the fate of mercury be of concern? How much concern? • Conclusion from these experiments: • decontamination works in these three model systems • methyl mercury can be reduced to elemental mercury

  17. BIOTECHNOLOGY: Phytoremediation Bioaccumulation of Arsenic • Arsenic is a major worldwide contaminant that can arise through • industrial activity (pesticides, mining, combustion etc.) • or from soil and ground water. • Associated with acute poisoning and linked to liver, lung, kidney, • bladder cancer; cause skin lesions; • damage to the nervous system. • Physical remediation (resins etc.) • Turn to bioremediation

  18. BIOTECHNOLOGY: Phytoremediation Bioaccumulation of Arsenic In India and Bangladesh (around the Bay of Bengal) ~400 million people are at risk of arsenic poisoning, and up to 40 million people drink well water containing toxic levels of arsenic.

  19. BIOTECHNOLOGY: Phytoremediation Bioaccumulation of Arsenic: General Strategy (Meagher & Heaton, 2005)

  20. BIOTECHNOLOGY: Phytoremediation Bioaccumulation of Arsenic: data from Dhankher, et al. (2002) • Strategy behind cloning • bacterial arsenate reductase (ArsC) catalyzes reduction of arsenate • to arsenite. • bacterial γ-glutamylcysteine synthetase ( γ-ECS) catalyzes the formation • of γ-glutamylcysteine (γ-EC) from glutamate and cysteine for synthesis • of glutathione (GSH) and phytochelatins (PCs; three arrows) • reduced arsenite can bind organic thiols (RS) such as those in γ-EC, GSH, • and PCs. Then transfer to vacuole.

  21. BIOTECHNOLOGY: Phytoremediation Bioaccumulation of Arsenic: data from Dhankher, et al. (2002) • Selection of SRS1p/ArsC9 (ArsC9) + ACT2p/ γ-ECS (ECS1) plants on 250μM arsenate. • Arabidopsis lines overexpressing ArsC and γ-ECS (ArsC9 + ECS1 or ArsC9 + ECS10). • Doubly transformed lines expressing both ArsC and γ-ECS(ArsC9 + ECS1 and • ArsC9 + ECS10), with parental lines SRS1p/ArsC9 and ACT2p/ECS1 line (ECS1) • grown on 200μM arsenate for four weeks. • ArsC is expressed from rubisco SRS1p small-subunit promoter and γ-ECS from the actin • ACT2pt promoter and terminator.

  22. BIOTECHNOLOGY: Phytoremediation Bioaccumulation of Arsenic: Possible metabolism (Doucleff and Terry,2002) Can further engineer plants to move the arsenic into leaf tissue so as to aid harvesting!!!

  23. BIOTECHNOLOGY: Phytoremediation • Phytoremediation of toxic compounds: • Explosives • Organomercury • Arsenic • Heavy metals such as Al, Cd, Cr, Cu, Fe, Mn, Mo, Ni, Pb, Se, V, and W. • ( plants extract B, Cl, Cu, Ca, Fe, K, Mg, Mn, Mo, N, Ni, P, S, Se, Zn from • the soil so there is great potential) • Mixtures of compounds??

  24. BIOTECHNOLOGY: Phytoremediation • Future of Plant Bioremediation: • Future cloning strategies • What plants should we use? • (annuals, trees prior to flowering, sterile?) • Why not use native plants? • Bioremediation of air? • Barriers to implementation • How to judge precautionary principle? • What are alternatives (resins) for water? • Costs of doing vs not doing?

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