Genetic modification in environmental

remediation

GENETIC
MODIFICATIONS
Sourav Mahajan
M.Sc. 3rd sem
AIMT
LET’S WATCH SOME PAST
AND PRESENT CONDITIONS
OF OUR MOTHER EARTH.
PRESENT SENARIO OF THIS WORLD
WORSE!!!

In 2009, near Krichim, Bulgarial, volunteers clean up discarded

plastic bottles and garbage clogging up the dam.
Antarctic penguins on ice floes. Global warming has threatened the
survival of the polar animals.
Iceberg in Antarctica. Affected by global warming, the two poles are
beginning to melt.
Brazil, a nut tree forest destroyed by a large fire.
The almost dried-up Curuai Lake in Brazil.
Comparison pictures of Everest glaciers. The above photo is the
Rongbuk Glacier during an analysis of Everest in 1968 by the Chinese
Academy of Sciences; The bottom photo was taken by Greenpeace in
2007 of the Rongbuk Glacier.
Comparison pictures of the Heyuan glacial lake. The
above picture was taken by Greenpeace’s John Novis in
June of 2005; The bottom picture was taken by
Greenpeace’s Du Jiang in September of 2005.
2009 December 6, Jammu, India: A person searches for recyclable
materials inside the Tawi River.
Tuvalu: Ocean waters flood a hole dug for
coral. Data shows that the country of Tuvalu
will be submerged by the ocean in 50 years.
In a fish farm in Linzhang Village on the outskirts of Changzhi
City of Shanxi Province, husband and wife fish farmers from
Fujian province Cheng Daocheng and Cheng Jinlan reluctantly
dispose of the catfish that have frozen to death. China News
Agency (Fao Yaolin photo)
Tourists sit on the open-air cafe tables, the chairs having already been
submerged by flood waters. November 30, Venice, Italy, was again
submerged by flood waters influenced by tropical storm tides, with the
water level reaching as high as 1.31 meters in certain areas. Photos from:
China News Web
2008 November 24, Yichang City of Hubei province, two man-made
culverts continuously “vomit” ink-like sewage into the Yangzte River,
dyeing the river water black. China News Service (Pu Xinyang photo)
September 23, dust storms occurred in the eastern region of Australia
that includes Sydney, affecting the local traffic. With the entire city of
Sydney enveloped in an orange cloud of dust, and owing to very low
visibility, airplanes and ferries were temporarily grounded or
suspended.
WHAT WILL BE THE FUTURE THEN??

Don’t ask the planet what’s

wrong, we need to ask ourselves!
LETS GET BACK TO OUR
STRATEGIES TO CURB
THIS DEVIL
 THREE STRATEGIES ARE WORKING CONJOINTLY-

1. Through chemical means

2. Through biological means
3. General awareness
Remediation by using living machines!!
 There are 3 mode of biological remediation
1. Through natural action of microbes-bioremediation.

2. Through natural action of some plants-

phytoremediation.
3.Through genetic modifications in microbes and in plants.
Phytoremediation
 Phytoremediation describes the treatment of
environmental problems (bioremediation)
through the use of plants which mitigate the
environmental problem without the need to
excavate the contaminant material and
dispose of it elsewhere.
 Plants can break down, or degrade, organic

pollutants or stabilize metal contaminants by

acting as filters or traps.
PHYTOREMEDIATION
Bioremediation

 Bioremediation or bioaugmentation is the

process of using naturally occurring microbes
to digest and convert unwanted waste
material into harmless substances.
MIGHTY MICROBES
 Deinococcus radiodurans bacteria have been
genetically modified to digest solvents and heavy
metals, as well as toluene and ionic mercury from
highly radioactive nuclear waste.
 Geobacter sufurreducens bacteria can turn uranium
dissolved in groundwater into a non-soluble,
collectable form.
 Dehalococcoides ethenogenes - The bacteria are
naturally found in both soiland water and are able
to digest the solvents much faster than using
traditional clean up methods.
MIGHTY MICROBES
 An enzyme from a bacterium, Thermus
brockianus, found in Yellowstone National
Park, breaks down hydrogen peroxide 80,000
times faster than current chemicals in use.
 The bacterium, Alcaligenes eutrophus,

naturally degrades 2,4-D, the third most

widely used herbicide in the U.S.
GENETIC MODIFICATIONS
 GENETIC MODIFICATIONS IN MICROBE
 GENETIC MODIFICATION IN PLANTS
 PRODUCTS OF MICROBES LIKE

NANOPARTICLES AND PHB GRANULES

GENETIC MODIFICATION IN MICROBES
 ALREADY KNOWN BIOREMEDIATING
BACTERIA-
 Pseudomonas, Alcaligenes.
 A NEW ENTRY IN THIS GROUP NAMED AS

Shewanella. August 31, 2009

 The team of researchers from the Georgia

Institute of Technology, Michigan State

University and the Pacific Northwest National
Laboratory analyzed the gene sequences,
proteins expressed and physiology of 10
strains of Shewanella.
Shewanella STRAINS.
SHEWANELLA
 Similar to a human breathing in oxygen and
exhaling carbon dioxide,
many Shewanella microbes have the ability to
“inhale” certain metals and compounds and
convert them to an altered state, which is
typically much less toxic.
 This ability makes Shewanella very important

for the environment and bioremediation, but

selecting the best strain for a particular
project has been a challenge.
SHEWANELLA
 If the strain is put in an environment that
contains high concentrations of uranium, that
microbe is likely to acquire the genes that
accept uranium from a nearby strain, in turn
preventing uranium from spreading as the
groundwater flows.
 This adaptability of bacteria is remarkable.
Part of genetic modification
 Konstantinidis’ team at Georgia Tech is
currently investigating communities of
these Shewanella strains in their natural
environments to advance understanding of
the influence of the environment on the
evolution of the bacterial genome and
identify the key genes in the genome that
respond to specific environmental stimuli or
conditions, such as the presence of heavy
metals.
Research paper
 Genetic Engineering of Escherichia
coli for Enhanced Uptake and
Bioaccumulation of Mercury
Weon Bae, Rajesh K. Mehra, Ashok Mulchandani, and
Wilfred Chen
Department of Chemical and Environmental
Engineering and Environmental Toxicology Program,
University of California, Riverside, California 92521
 Received April 25, 2001; Accepted September 6, 2001.
Genetic modifications in plants
genetic engineering in Eastern cottonwood tree (Populus
deltoides)
 Researchers Richard Meagher, from UGA's
Department of Genetics, and Scott Merkle, from
UGA's Warnell School of Forestry, used genetic
engineering to produce an Eastern cottonwood tree
(Populus deltoides) capable of cleaning up
contaminated soil through a process known
as phytoremediation.
 In a study published in 2007, Meagher and Scott
describe their engineering of cottonwoods to
extract toxic mercury from the soil.
 The trees then release the vapor into the
atmosphere, where it is diluted to harmless levels.
Phytoremediation
 Major successes have also been achieved in
producing the biodegradable plastic polymer
polyhydroxybutyrate (PHB) in Arabidopsis.
 Nylon and polyester-like compounds are not

far behind!!!
 Plants have a remarkable ability to extract

and concentrate elements and compounds

from air, water, and soil.
Mercury breathing plants
 Bacterial mercuric ion reductase, MerA, is
effective in catalyzing the reduction of
mercuric ion, to elemental mercury by using
NADPH as an electron donor .
 MerA is a soluble, NADPH-dependent, FAD-

containing, disulfide oxidoreductase.

 Rugh et al. began this fascinating research

project with the goal of increasing mercury

resistance and volatilization in plants
expressing merA.
What is the difficulty??
 The difficulty was that the bacterial merA
gene was heavily skewed toward GC-rich
sequences, contained no plant translation
signals.
 To ensure translation in plants, the authors

built a synthetic gene, merApe9, with a

lowered GC composition and few rare codons
and CpG dinucleotide.
Mercury breathing plants
 In addition, the 5' region immediately
upstream of the initiator ATG codon was
replaced with consensus plant and
Escherichia coli translation signals.
 The modified merApe9 encoded a functional

mercuric ion reductase in transformed E. coli.

 Agrobacterium tumefaciens-assisted

transformation was used to introduce

merApe9 into the recipient plant, Arabidopsis
thaliana.
Result
 Analysis of T2 and later generation plants
showed greater resistance to germination
and growth on medium containing HgCl2
concentrations of 25-100 µM.
 These levels were toxic to most plant species

but not to the transformed Arabidopsis.

 Rapid evolution of mercury vapor by relative

to control plants confirmed resistance was

due to mercury reduction.
 Plants and bacteria expressing merApe9

-more resistant to toxic levels of Au2+ also.

Future works
 The efficiency of phytoremediation can be
improved by expressing high-affinity, high
selectivity metal-binding peptides interand
extracellularly.
 Well-known metal binding proteins include

metallothioneins, metalloenzymes; metal-

activated enzymes; and various metal
storage, carrier, and channel proteins.
 They are widely distributed in yeast and in

lower and higher plants.

Future works

 Metal binding domains from newly discovered or already
known proteins may be incorporated in synthetic genes
encoding low molecular weight peptides, which can be
expressed in plant roots, the vascular system, and, if
necessary, shoots.
Future works
 In addition, sequential rounds of randomized
mutagenesis followed by selection under increasing
metal binding stringency can be used to
modify and improve metal binding efficiency
and selectivity of known and newly identified
metal binding proteins.
Future works
 Genes that encode such high-affinity
transporters may be used to engineer plants
capable of enhanced accumulation of
particular metals.
A recent patented product-ECOBIOTIX
Remedy of all!!!
 This solution in a non-hazardous/non-
corrosive remediation agent.
 It is 100% green chemistry, an enzyme that is

non-hazardous, non-corrosive and non-toxic

and is biodegradable. 
 It neutralizes soil toxicity to support healthy

plant growth and makes it ideal for open

water applications
A recent patented product-ECOBIOTIX
Remedy of all!!!

 This enzyme is now being used by the U.S.

Supreme Court to rid the toxin and smell
within the lines of the sewers. 
 It has also been used to clean off

contaminated equipment from the Iraq war.

LETS CHECK OUT THIS GOD THING!!!

VIDEO OF ECOBIOTIX
"What's the use of a fine house if
you haven't got a tolerable planet
to put it on?"
- Henry David Thoreau

LET’S TAKE A SERIOUS ACTION MY

FRIENDS TO SAVE OUR MOTHER
EARTH!!!
GREENPEACE ACTION
 REFERENCES

ARTICLES FOR PHYTOREMEDIATION

 Fox, B. & Walsh, C. T. (1982) J. Biol.Chem. 253, 4341-

4088.
 Lehman, N. & Joyce, G. F. (1993) Nature (London)
361, 182-185.
 Kumar, P. B. A. N., Dushenkov, V., Motto, H. &
Raskin, I. (1995) Environ. Sci. Technol.29, 1232
 Dushenkov, V., Kumar, P. B. A. N.,Motto, H. & Raskin,
I. (1995) Environ.Sci. Technol. 29, 1239-1245.
 REFERENCES
ARTICLES FOR BIOREMEDIATION BY MICROBES

 Alonso A., Rojo, F. and Martinez, J.L. (1999) Environmental and

clinical isolates of Pseudomonas aeruginosa show pathogenic
and biodegradative properties irrespective of their
origin. Environ. Microbiol., 1, 421–430. [PubMed]
 Cases I. and de Lorenzo, V. (2001) The black cat/white cat

principle of signal integration in bacterial promoters.  EMBO

J., 20, 1–11. [PMC free article] [PubMed]
 Harvey S., Elashvili, I., Valdes, J.J., Kamely, D. and Chakrabarty,

A.M. (1990) Enhanced removal of Exxon Valdez spilled oil from

Alaskan gravel by a microbial surfactant.  Biotechnology, 8, 228–
230. [PubMed]

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