Issues in Biotechnology:

The Way We Work With Life

Dr. Albert P. Kausch

life edu.org
Section II

The Applications of
Biotechnology
A Sweeping General Survey on Life and Biotechnology
The University of Rhode Island

Agricultural Biotechnology
life_edu

Lectures 13 and 14

Issues in Biotechnology:
The Way We Work With Life
Dr. Albert P. Kausch

life edu.org

Agricultural Biotechnology
Part I Where Does Our Food Come From?
Part II DNA-based Biotechnology
And Modern Agriculture
Part III. Issues, Controversies and Concerns
a. Setting the Stage about Food and Agriculture:
b. Issues, Controversies and Concerns
c. The Organic Food Debate
Part IV. The Ethics
of Agriculture
life_edu
Part V. Renewable Energy and the Future of Humanity

Issues in Biotechnology:
The Way We Work With Life
Dr. Albert P. Kausch

life edu.org

Agricultural Biotechnology
Part I Where does our Food Come From?

life_edu

Issues in Biotechnology:
The Way We Work With Life
Dr. Albert P. Kausch

life edu.org

Agricultural Biotechnology
Part II DNA-based Biotechnology
And Modern Agriculture
Lectures 11 and 12

life_edu

Issues in Biotechnology:
The Way We Work With Life
Dr. Albert P. Kausch

life edu.org

Agricultural Biotechnology
Part III. Issues, Controversies and Concerns
Setting the Stage about Food and Agriculture:

life_edu

Do you eat genetically modified (GM) foods?

(A)Yes
(B) No
(C)Sometimes
(D) I dont know
(E) I dont care

Ifyes,whatfoodsoringredientsdidyou
avoidoreatlessof?

Foods/ingredientsavoided
A.Sugar/Carbohydrates
B.Fats/Cholesterol
C.AnimalProducts
D.Salt/Spices
E.Biotechnologyproducts

life_edu

its not biotech!

Ifyes,whatfoodsoringredientsdidyouavoidoreatlessof?
(Openended;Multipleresponsesallowed,n=478)
Foods/ingredientsavoided

3/05

Sugar/Carbohydrates

58%

Fats/Cholesterol

37%

AnimalProducts

34%

Salt/Spices

14%

SnackFoods

11%

Biotechnology

<%
life_edu

IFIC

Agricultural Biotechnology
What is it?
Where does our food come from?
History of Agriculture
Plant Domestication
How is DNA-based biotechnology used for crop
improvement?
How is it done?
What are the goals?
What as been done so far?
What is in the future?
What are the controversies
life_edu and concerns?

Wild Plants
Crop Plants
Genetics
Breeding
Heredity
DNA

Conventional Foods
Processed Foods
Organic Foods
Natural Foods
Whole Foods

Crop Plants
Conventional farming
Organic farming

Past-Present-Future
Conventional
Breeding

Biotechnology

life_edu

Issues in Biotechnology
Applications in Biotechnology
Agricultural Biotechnology

life_edu

Agricultural Biotechnology
Genetically Modified Foods:

Panacea
Or Pandoras Box?

life_edu

Agricultural Biotechnology
Genetically Modified Organisms GMO
How is it done?

It is now possible to
clone any gene
from any organism
and move it into
plants
life_edu

Issues in Biotechnology
The Way We Work With Life

II. Applications in Biotechnology

Dr. Albert Kausch

GMOs in Food:
Risks and Benefits

life_edu
Lecture 14: Agricultural Biotechnology: GMOs in Food Risks and Benefits

It is now possible to clone

any gene from any
organism and move it
into plants

life_edu

Plasmids are circular pieces

of DNA found in some bacteria
Many copies per cell
Antibiotic resistance gene
Plasmids can be cut and
pasted back together
Foreign genes can be
inserted
life_edu

How is a gene cloned?

Boyer, Cohen, and Berg, 1972

Enzymes were discovered

that cut DNA
at specific sequences
And subsequently,
enzymes were discovered
that paste DNA together.
The ability to cut and
paste DNA
allowed gene cloning.

life_edu

How is a gene cloned?

Foreign DNA (gene)
is inserted into a plasmid
that has a gene for
antibiotic resistance
The plasmid is introduced into
a bacterial cell and grown on
the antibiotic

Only bacteria with the plasmid

growthe inserted gene is
life_edu
copied many times.

Anatomy of a Transgene

Promoter

Coding Sequence

Terminator

Protein coding sequence

Cell specificity
Developmental specificity
Start transcription

Stop transcription
Message stability

Gene constructs can be moved into plants and the gene is expressed
driven by the promoter sequence
life_edu

 life_edu

Essential Components for

Successful Plant Transformation
Tissue Culture System *
regeneration of whole plants from single cells

Gene Construct
Molecular vector construct using foreign DNA

Method for DNA Delivery

Agrobacterium tumefaciens
Direct DNA Uptake
Whiskers
Sonication
Microprojectile Bombardment

Efficient Selection Strategy

differentiation between cells which receive the

transgene from those that dont
Antibiotic Resistance
life_eduResistance
Herbicide

Cloning of Plants: Plants can be grown back from a

single cell this ability is called totipotency.
Steward 1958 Cornell Univ.

Many, but not all plant cells are totipotent.

Tissue culture is a key element for creating
transgenic plants. life_edu

Regenerable Cell Culture Systems

Whole plants can be
regenerated back
from single cells or
groups of cells

life_edu

Essential Components for Successful

Gene Transfer to Plants
Tissue Culture System
regeneration of whole plants from single cells

Gene Constructs: Using DNA cloning

Molecular vector construct using foreign DNA

Method for DNA Delivery

Agrobacterium tumefaciens
Direct DNA Uptake
Whiskers
Sonication
Microprojectile Bombardment

Efficient Selection Strategy

differentiation between cells which receive the

transgene from those that dont
Antibiotic Resistance
life_eduResistance
Herbicide

Transgenes confer new traits

Promoter

Coding Sequence

Terminator

Your favorite gene

Herbicide resistance
Controlled expression
Stop transcription
Pest resistance
on in only specific cells
Message stability
insect, fungus, virus
Drought, salt,
and freeze tolerance
Nutritional enhancement
proteins, vitamins, oils
Specialty chemicals
life_edu
pharmaceuticals, bioplastics

How are cloned genes moved into cells?

Microinjection
Chemical treatments
Bombardment

Electroporation
Microprojectile
life_edu

Early work on Plant transformation:

Laboratory of Marc Van Montague, Gent, Belgium
Agrobacterium tumefaciens: Gene transfer to plants

life_edu

Agrobacterium tumefaciens:
a gene transfer vector for plants
A naturally occurring soil bacterium
which transfers DNA to plants.
Allowed the first genetically
engineered plant (tobacco) 1983
Used to transfer genes to dicot
plants, however not readily amenable
to cereal crops
life_edu

The Gene Gun:

Invented by John Sanford (1986)

life_edu

Particles shot by the gene gun enter the cell and deliver DNA

life_edu

DuPont bought the rights

to Sanfords device

life_edu

BIO-RAD Biolistic PDS-1000/He Particle Delivery System

Allows testable and
reproducible parameters:
Gap distance
Rupture discs
Macrocarrier flt distance
Particle flt distance
Particle size
Particle density
Target cell positioning
Cell/Tissue pre-treatments:
osmotic
silver nitrate
numerous others

life_edu

Agrobacterium is now
a vector for monocots
and dicots
Agrobacterium and
the gene gun
are viable approaches to
plant gene transfer

life_edu

Essential Components for Successful

Gene Transfer to Plants
Tissue Culture System
regeneration of whole plants from single cells

Gene Constructs: Using DNA cloning

Molecular vector construct using foreign DNA

Method for DNA Delivery

Agrobacterium tumefaciens
Direct DNA Uptake
Whiskers
Sonication
Microprojectile Bombardment

Efficient Selection Strategy

differentiation between cells which receive the

transgene from those that dont
Antibiotic Resistance
life_eduResistance
Herbicide

A Selectable Maker Gene

A method for selecting cells that have received the DNA from those
that have not.

Promoter

Constitutive expression
on in all cells

Coding Sequence
Selectable marker gene
Herbicide resistance
Antibiotic resistance

life_edu

Terminator

Stop transcription
Message stability

Selectable marker genes allow transformed cells

to be recovered from a non-transformed
population

Herbicide resistance genes were among the first selectable markers

life_edu

Transformable & Regenerable

Cell Culture Systems
Selected
transformed cells
can be regenerated
back
to whole plants all
of which carry the
new gene.

life_edu

The First Genetically

Engineered Corn
Herbicide Resistance
Roundup Ready
DeKalb Genetics
Corporation

life_edu

Now over 95% of US

corn is genetically
modified

The process of producing transgenic corn requires

9 months to a year to recover seed with a new gene

life_edu

The point is that an inserted foreign gene is transcribed

into RNA and translated into protein just like any gene.

DNA
RNA
Protein
A foreign gene will produce a new
protein that may confer new traits
life_edu

Issues in Biotechnology
Gene constructs can be readily made in the laboratory by
fusing together DNA segments from different sources.
If done correctly, these cloned constructs can be
delivered into plants where there will be stably
integrated into the plants DNA and expressed to confer
new and useful traits. Gene constructs can be
introduced into plants by
(A) microprojectile bombardment (also known as the gene
gun or biolistics) delivers DNA by coating small particles
of gold with the vector and shooting them into plant cells
(B) all of the examples shown have worked
(C) Microinjection: by using a small needle to inject DNA
directly into plant cells
(D) Agrobacterium tumefaciens:alife_edu
bacteria that acts as a
natural gene transfer vector to deliver DNA into plants

It is now possible to move

a single gene or genes into
a known genetic
background.

life_edu

The
Importance
of Reporter
Gene
Constructs
Transgenic plants
have been made
which express
firefly luciferase
And Green life_edu

fluorescent protein
Bioluminescence Green
from Aequorea victoria
APPLICATIONS
Transgenic research: watching for
expression
A unique reporter gene
And many more uses!!!
Martin Chalfee wins the Nobel
Prize for GFP!

life_edu

Clone of the Week

GFP fluorist A GFP modified daisy

Istituto Sperimentale per la Floricoltura in San Remo, Italy
life_edu

Promoter analysis for gene expression

in transgenic maize
Promoter
pep carboxlyase
adh1
CaMV 35S
aldolase 1
actin 1
endosperm specific

Reporter gene
GUS
GFP
C1B
LUX

Assay
Stable transformants
Histological
mRNA in situ
Enzymatic assays
PCR
Southern blots

Endosperm-specific GUS expression Adh1 GUS in hypoxia

induced maize seedlings
shown as days post-pollination

life_edu

CaMV 35S-GUS

Transgenic Corn expressing a bacterial gene, GUS.

A promoter that turns on

gene expression in all
plant cells was used to drive
expression of a coding
region for a gene
(beta-glucuronidase or GUS)
from a bacteria (E. coli)
in corn. Expression of this
marker gene is visualized by
a histological reaction
that stains the cells blue.

Reporter genes, like GUS

allow specificity of the
promoter to be seen.

life_edu

Cloned DNA can be inserted

into the chromosomes of plants.

life_edu

Gene transfer from one organism to another is not n

Image of two species of
bacteria transferring
viral
phage particles
Bacteria transfer genes
to other bacteria and
plants.

Now in nature there

is another organism
capable of
Transferring DNA:
we call that organism
a human being.

life_edu

Traits and Genes

Second and additional trait genes are piggy-backed in
and are co-expressed with the selectable marker gene.
Thus, two genes (and more) can be delivered at the same time
In addition, genes can be stack by crossing transgenic plants

life_edu

Trait Modification for

Agricultural Crop Improvement
Past and current traits in transgenic products on the market
Traits in development and the future

life_edu

Genetically Modified Organisms

Transgenes confer new traits
Promoter

Coding Sequence

Terminator

Your favorite gene

Herbicide resistance
Controlled expression
Stop transcription
Pest resistance
on in only specific cells
Message stability
insect, fungus, virus
Drought, salt,
and freeze tolerance
Nutritional enhancement
proteins, vitamins, oils
Specialty chemicals
life_edu
pharmaceuticals, bioplastics

Domestication of the Wild Tomato

life_edu
.And
Genetic Modification

The first commercially available

genetically engineered plant is released, (1992).
Made using Agrobacterium.

life_edu

Herbicide tolerance
Herbicide tolerant crops allow farmers to apply
a specific herbicide to control weeds without harm to the
crop. Gives farmers greater flexibility in pest management
and promotes conservation tillage.

Now: soybeans, cotton, corn, canola, rice

Future: wheat, sugar beet
life_edu

Insect Resistance

Bt crops are protected against insect damage and reduce

pesticide use. Plants produce a protein -- toxic only to certain
insects -- found in a common soil bacterium called Bacillus
thuringiensis, or Bt.

Now: corn, cotton, potatoes

Future: sunflower, soybeans,
canola, wheat, tomatoes
life_edu

Transgenic Corn
Corn with a gene from a bacteria, Bacillus thuringiensis, which codes for a protein
that is toxic to lepidopteran insects

life_edu

Corn expressing the Bt gene

Provides insect resistance
to European Core Borer
without the use of insecticides
millions of acres in the US
already in consumer foods
Monsanto
life_edu

What,ifanything,areyoumostconcernedabout
whenitcomestofoodsafety?
FoodSafetyConcerns
A.Handling/Preparation
B.Disease/Contamination(foodborneillness)
C.Ingredients(chemicals,preservatives,fats)
D.Chemicals/Pesticides
E.GMOsinfood

<

life_edu

Biotech not a top-of-mind food safety

concern
What,ifanything,areyoumostconcernedaboutwhenitcomestofoodsafety?
(Open-ended; Multiple responses allowed, n = 1000)

FoodSafetyConcerns

3/05

Handling/Preparation

42%

Disease/Contamination(foodborneillness)

28%

Ingredients(chemicals,preservatives,fats)

23%

Packaging(expirationdates,packaging)

14%

Chemicals/Pesticides

7%

Biotechnology
life_edu

<%
IFIC

Fungus Resistance
Chitinase
Glucanase
Lytic Peptides
Other Anti-Fungal
compounds

life_edu

Oils

High-performance cooking oils

maintain texture at high
temperatures, reduce need for
processing and create healthier
food products. The oils are either
high oleic or low linolenic. In the
future, high stearate.
Future: sunflower, peanuts and
soybeans
Healthier cooking oils have reduced saturated fat.
Future: soybeans
life_edu

Nutritionally enhanced foods

Golden Rice
Nutricueticals: Nutritionally enhanced
foods will offer increased levels of nutrients,
vitamins and other healthful
phytochemicals. Benefits range from helping
developing nations meet basic dietary
requirements to boosting disease- fighting
and health-promoting foods.

Future: protein-enhanced sweet potatoes and rice; high-vitaminA canola oil; increased antioxidant fruits and vegetables.
life_edu

How to Make
Golden Rice

3.The Agrobacterium are then added to

1. The genes that give golden rice
rice cell cultures. The Agrobacterium transfers
its ability to make beta-carotene
the engineered genes that encode
in its endosperm come from
theinstructions
daffodils and a bacterium,
for making beta-carotene into the rice cells.
Erwinia uredovora
2. These genes along with
promoters (segments of DNA that
activate genes) are inserted into
plasmids (small loops of DNA) that
occur inside a Agrobacterium
vector

4. The rice cells are grown back to whole

plants carrying the new genes. These
transgenic plants can be crossed to rice
suited to particular regions around the world.

life_edu

 life_edu

How Could Anyone Disagree?

Furthemore, Dr. Potrykus violated seventy five

international patents to accomplish this task.
Perhaps he should be thrown in jail if it is distributed?
life_edu

Issues in Biotechnology

The first commercially available genetically engineered

plant was released in 1992. It was
(A) a slow ripening tomato called the Flavr Savr
(B) a glow in the dark daisy with a GFP protein from
fluorescent jellyfish
(C) high vitamin A rice called Golden Rice
(D) a corn plant with resistance to the herbicide Round-Up

life_edu

Fruit and Vegetable Quality

Delayed-ripening fruits and
vegetables have superior flavor, color
and texture, are firmer for shipping and
stay fresh longer.

Now: tomatoes
Future: raspberries, strawberries, cherries
tomatoes, bananas, pineapples

life_edu

Edible Vaccines in
Transgenic Plants
Antibodies produced in Plants
Specialty Chemicals

life_edu

Biofuels: A Sustainable Solution

Dr. Albert P. Kausch
University of Rhode Island

life

edu.org

Genetically Modified
Plants for Biofuels-Some
Considerations:
Thinking Outside the
The Role of
Barrel
Agricultural
Biotechnology
life_edu

The Genetic Engineering

Yield

Yield
Of Many Genes

life_edu

Controlling Complex Traits

Yield

Yield

Quantitative Trait Loci

life_edu

Yield Stability for Crops

Abiotic Stress Tolerance

Drought
Freeze
Salt

Improving stress
tolerance in
plants by gene
transfer
osmolytes
osmoprotectants
membrane fatty
acids

life_edu

Heat
Chilling
Flooding

 life_edu

Stress tolerant maize at the beach

life_edu

Wild Type Maize

life_edu

The Future of Plant Biotechnology

Edible Vaccines
Phytoremediation

High Protein Content

Drought Tolerance

Disease Resistance

Pest Resistance life_edu

Salt Tolerance

Cold Hardiness

Isthereanyinformationnotcurrentlyon
foodlabelsthatyouwouldliketoseeadded?

A.
B.
C.
D.
E.

No
Ingredients
Nutrition
GMOs
Other (i.e. safety and handling)

life_edu

Biotech labeling
not a top-of-mind consumer demand
Anyinfonotcurrentlyonfoodlabelsthatyouwouldliketoseeadded?
(Open-ended; Multiple responses allowed, n = 1000)

life_edu

IFIC

DoyoutrusttheFDA?
A.
B.
C.
D.
E.

Trust
Dont trust
Somewhat trust
Somewhat distrust
Dont know/Refuse to answer

life_edu

FDArequiresspeciallabelingwhenafoodis
producedundercertainconditions:
Speciallabelingisnotrequired.Would
yousaythatyousupportoropposethis
policyofFDA?
A.
B.
C.
D.
E.

Support
Oppose
Neither support or oppose
Dont know
Refuse to answer

life_edu

7 . While it has not yet been done for all organisms on earth,
based on what we now know, it is possible to clone any gene
from almost any living organism, transfer that gene into
another unrelated organism, and make the same protein coded
for by that gene. This means that the genetic code is a
universal language for life on this planet.
a. this statement is generally true
b. this statement is generally false
c. there is still not enough information
d. this is confidential information held only by the government
e. it is therefore impossible to use this information to create
new plant cultivars

8 . Former President Jimmy Carter has said that

Biotechnology is not a threatstarvation is. Vitamin A
deficiencies are responsible for a million lives a year globally
and many of those are in developing countries where the main
staple is rice. Dr. Ingo Potykus of Switzerland genetically
modified rice to accumulate high levels of beta carotene the
natural precursor for vitamin A. Some have argued that it
would be unethical to deny this population of such a useful tool
to alleviate suffering. This variety is known as:
a. Terminator rice
b. Golden rice
c. Fried rice
d. Green Revolution rice
e. The Gold Revolution

9. It is interesting that because all life on this planet uses

information in the form of either DNA, RNA or both, and that
the same bases are used, and that the genetic code is almost
always the same using three of those bases to code for one of
twenty amino acidsbecause now genes can be cloned and
moved from one organism into another and the foreign gene
will make the same protein! So a gene from a bacteria can be
expressed in plant to make the same protein that was made in
the bacteria. The process of taking a gene from one species
and expressing it in the genome of another species is called
a. transgenics
b. eugenics
c. carcinogenics
d. pharmacogenomics
e. agricultural forensics

10. The first commercially available genetically engineered

plant was released in 1992. It was
a. a slow ripening tomato called the Flavr Savr
b. a glow in the dark daisy with a GFP protein from
fluorescent jellyfish
c. high vitamin A rice called Golden Rice
d. a corn plant with resistance to the herbicide Round-Up
e. a seedless watermelon made by radiation

11. Gene constructs can be readily made in the laboratory by

fusing together DNA segments from different sources. If done
correctly, these cloned constructs can be delivered into plants
where there will be stably integrated into the plants DNA and
expressed to confer new and useful traits. Gene constructs can
be introduced into plants by
a. microprojectile bombardment (also known as the gene gun
or biolistics) delivers DNA by coating small particles of gold
with the vector and shooting them into plant cells
b. all of the examples shown have worked
c. Microinjection: by using a small needle to inject DNA
directly into plant cells
d. Agrobacterium tumefaciens: a bacteria that acts as a
natural gene transfer vector to deliver DNA into plants
e. osmotic or electric shock treatments

12. It is now possible to make gene constructs in the laboratory

by fusing together DNA segments from different sources. If
done properly, these cloned constructs can be expressed in
plants to confer new and useful traits. For example, genes that
encode proteins from a bacteria can be moved into plants to
make them resistant to pests without using pesticides. This
process of plant improvement has been
a. only theoretical but hotly debated for over ten years
b. only accomplished by big companies but not by academic
labs
c. applied widely and successfully in US agriculture for over
fifteen years
d. has been shown to be fraudulent and dangerous to people
and livestock
e. shown to be a link in the rising incidence of autism in the US

13. Agricultural Biotechnology:

a. is a new form of organic agriculture that uses synthetic
manure with the promise of providing safer and more healthy
plants
b. includes domestication, varietal selection, genetics, wide
crosses, mutagenesis, and gene transfer for the overall goal to
help produce new varieties of crop plants that are useful for
people
c. a unnatural process for developing genetically modified
(GM) plants that have been shown to cause birth defects in
humans
d. is too recent of a technology to be of practical use, but has
great promise for the future to aid in global food resources
e. has now been banned by all US farmers and grocery sores
as inherently unsafe

14. GMO is
(A) a designation used by a group called Germplasm
Movement from Origins that tracks the origins of first crops
domesticated by humans ~10,000 yrs ago
(B) . a designation for 'Genetically Modified Organism'
indicated the use of transgenics
(C). the acronym for militant Global Motivation
Organization, a non-governmental organization that tracks
the release of genetically engineered plants as something they
think is harmful to the environment, health and safety of all
US citizens
(D). a crop that is 'Grown More Organically' as a designation
of safety used by the USDA for crops that have a clear
environmental and health advantage
(E). a type of IED developed by the US specifically for use in
Iraq and Afghanistan to control the production of Opium
poppies

Next.Agricultural
Biotechnology:
Part III:
Issues, Controversies and
Concerns