BSCI124 Lecture Notes
Undergraduate Program in Plant Biology, University of Maryland
LECTURE 41 - PLANT IMPROVEMENTS: BIOTECHNOLOGY
I. Plant Breeding (in Maryland - 100 genetic companies)
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A. Traditional method of improving plants - selective breeding
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1. Selection of desired characteristics: yield, palatability, resistance
to disease and insects, aesthetic characters
2. Hybridization - bringing together desired genes (by controlled mating)
from two or more individuals; results in a combination of desired characters
in the offspring - a hybrid
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a. farmers would save seed from the best plants to grow next year
B. Green Revolution
- The example of successful crop improvement (dwarf, high yielding wheat
varieties)
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1. Norman
Borlaug, awarded the Nobel Prize in 1970 for his role in the Green
Revolution
2. Borlaug is remains concerned about the future noting in particular
the international agribusiness control of genetic material
C. Limitations
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1. Can only use genes
from within one species or several closely related species or wild species,
becoming a limitation due to loss of genetic diversity
2. Takes many years to develop an improved variety
II. Recombinant
DNA
[Important Illustration] and the new Genetics:
Genetic
Engineering
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A. Introduction
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1. In the early 1970's a Moratorium on a certain type of research was called
by those doing the work
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a. legislation was introduced before the US congress which would require
congressional approval for these experiments
b. involves a newly developed technology of gene
manipulation [Important Illustration]
c. experiments were deferred for 18 months so that an assessment could
be made regarding the potential danger of the research
2. Technique: ability to construct new combinations of DNA molecules, which
do not exist naturally= Recombinant DNA. Remember the Central
Dogma of Molecular Biology?
III. Combines two different technologies: Restriction Enzymes and
Bacterial Plasmids
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A. Restriction
Enzymes [Important Illustration]
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1. The real basis for the recombinant
DNA techniques [Important illustration]: Sequence specific DNases:
Recognize short sequences of bases in DNA, and make a double stranded cut
in the DNA molecule
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a. function in bacteria- destroy foreign DNA which might enter the cell
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1) each bacterium has its own restriction enzymes
b. each enzyme recognizes only one type of sequence
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1) sequence recognized is called a Palindrome
2) reads the same on the two strands in the opposite direction
3) example: G A A T T C
C T T A A G
some enzymes cut straight across, and others make
a staggered cut.
G A A T T C
C T T A A G
G AATTC
CTTAA G
Creates fragments of DNA, all with the same
overlapping ends.
2. Value of restriction enzymes
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a. each enzyme only recognizes the same palindrome regardless of the source
of DNA
b. so, fragments, from different sources, produced by the same enzyme,
contain the same overlapping ends
c. treat with ligase = permanently joined together = Recombinant DNA
molecule
B. Plasmids
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1. Extrachromosomal genetic elements in bacteria
2. Closed circular DNA molecules
3. Replicate independent of the chromosome = many copies/cell
4. Contain genes controlling such things as fertility, and antibiotic
resistance
C. Molecular cloning
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1. The usefulness of the technology combines plasmid biology with restriction
enzymes in the following way
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a. join a DNA fragment from one source to a bacterial plasmid which has
been cut once with the same restriction enzyme
b. plasmid is now called a vector and carries a foreign piece of DNA
c. recombinant plasmid is reintroduced back into a bacterial cell
d. inside the bacterial cell, the plasmid will carry one or more antibiotic
resistance factors, so that when plated on a medium containing the antibiotic,
only the strain of interest will grow. Plasmid is replicated and the number
increased greatly = MOLECULAR CLONING
IV. Actual gene transfer to plants: - what
is a transgenic plant
(see this-process
of making transgenic plant - animation
of process)
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A. Agrobacterium tumefaciens - a bacterium that causes crown gall
disease in many plants; unique among pathogens because it causes the disease
by transferring bacterial DNA from its plasmid into the plant's chromosome,
causes tumors
B. Tissue
culture - used to regenerate an entire plant from a single cell; involves
growing cells under sterile conditions with different nutrients and hormones;
in some plants you can grow entire new plant from one cell:
C. Plant
genetic engineering using Agrobacterium tumefaciens: method
to have one plant species (tobacco - easy to grow, we know a lot about
their breeding) express a gene from a different species
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1. Leaf disk
method of plant transformation
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a. punch out leaf disks
b. incubate with Agrobacterium culture carrying foreign gene
c. place disks on culture medium to induce regeneration and to select
for transgenic plants
d. root regenerated plants
e. every cell in the transgenic plant has the foreign gene
f. these plants pass genes on to their offspring
V. Applications of technology
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A. Getting a gene from one species into another and having it expressed
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1. Example - bacteria
that makes human insulin, insulin extracted from cows, but gene not
human/ now human gene in bacteria to produce insulin
2. Example - tobacco plant that glows in the dark (luciferin gene from
fireflies), transfer of gene not just from one species to another but from
one kingdom to another
B. Examples of application of gene technology in agriculture
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1. Herbicide
resistant plants
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a. in some areas, crops genetically modified for herbicide tolerance could
decrease the amount of herbicide used and allow for no-till agriculture,
which can minimize erosion
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1) Roundup
- a herbicide that is effective, non-toxic to animals, short-lived in environment
2) Research groups developed crop
plants resistant to Roundup, notably soybean, corn, tomato, potato,
wheat, cotton, alfalfa, etc.
2. Insect
resistant plants
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a. Bt toxin gene is cloned from bacteria and expressed in plants to provide
resistance from insect without the need for insecticides - data
about Bt toxin
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b. currently in crops of corn, corn, cotton
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c. current controversies
3. Controlled ripening tomato
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a. two varieties of tomato now on the market engineered for delayed ripening
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1) one tomato variety
has an extra gene - a reverse copy of the gene responsible for an enzyme
that breaks down cell walls; as a result, the
tomato softens more slowly
2) the other variety has a gene that controls the enzyme necessary
for the production of ethylene, one factor that makes a tomato soft
4. Enhanced resistance to viral diseases
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a. Crops bioengineered for pest resistance could increase yield, eliminate
the use of several insecticides now derived from fossil fuel, and reduce
health risks and groundwater contamination from pesticides
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1) used Agrobacterium to insert gene from viral protein into plant
cells
2) when regenerated, the plant (tobacco) produced this protein
and were more resistant to the virus
3) currently employed in
papaya and squash
5. Production
of vitamin A in rice
a. "golden rice" -a transgenic rice in which the genes for the production
of vitamin A have been inserted
(news
article)
6. Future
ideas
VI. Advantages of gene technology
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A. Foreign genes (from outside species) can be introduced
B. Potentially faster than traditional plant breeding
C. Specific genes can be transferred; much more control than in traditional
plant breeding
VII. Other
current and possible benefits of gene technology
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A. Already benefits in human health (human insulin).
B. Genetically engineered fish that grow much faster than wild (growth
hormone gene).
C. Recombinant bovine growth hormone already enables cows to use feed
more efficiently and produce more milk.
D. Gene Therapy: Use this technology of gene transfer to correct genetic
defects; any human diseases known to be due to a single gene defect; use
to correct single gene defect and cure disease
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1. Severe Combined Immune Deficiency. Bubble baby. due to a single gene
defect- adenosine deaminase
2. Lesch-Nyhan- severe mental retardation, self destructive behavior.
One brain enzyme missing
3. Cystic Fibrosis- single gene, inhale DNA with normal gene
E. The future of genetic research: One
review
X. Pros and cons of genetic engineering
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A. Cons- risks
and concerns
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1. Herbicide-resistance or insect-resistance genes could spread from the
engineered crops to wild relatives and create weeds that are especially
difficult
to control
2. Some scientists fear that the USDA does not require sufficient precautions
to prevent the spread
of genes from engineered plants to their wild relatives in field trials
3. Some bioengineered products could wipe out the major exports of
some developing nations
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a. example: a genetically altered bacterium is under development that produces
vanilla flavoring; this could eliminate markets for the vanilla beans,
one of Madagascar's major agricultural products
b. bovine growth hormone too expensive for small dairyman, so can't
compete with big companies
4. Biological control may solve the problem, cheaper and more effectively,
as shown with the work on the cassava
mealybug by Dr. Hans Herren, winner of the 1995 World Food Prize
B. Pros
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1. Almost 100 million people are expected to be added to the world's population
each year for the next 30 years
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a. some believe that without biotechnology, we won't be able to increase
the availability of affordable basic food.
b. although biotechnology has potential risks, starvation is worse
Other Sites of Interest:
All about Arabidopsis: "Guinea pig of the plant
world" by Bryan Ness
Biotechnology
and Agriculture
A
series of short reviews; excellent summaries!
Access Excellence: WWW based
teaching resources sponsored by Genentech, Inc.
Brief review
and excellent definitions
Bio Online: Links to biotech information
BioBlox: Biotechnology
Resource Center
A primer on molecular
genetics but pretty technical
BioEthics:
Links to resources on bioethics (little botanical)
Bt toxin links-
USDA National Agriculture Library
Code
of Conduct for Plant Biotechnology
A
discussion of plant breeding, still a bit technical
CIMMYT:
Where Norman Borlaug started the Green Revolution
Reviews
of
books and articles on the Green Revolution
Perils Amidst
the Promise: Ecological risks of transgenic crops in a global market
by Jane Rissler & Margaret Mellon -- An excellent review worthy of
serious review
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Last revised: 5/13/99 - Bottino, 12/2000 Straney