Plant engineering

1
Plant engineering

• The perpetual search for improvement

2
Salute to Norman E. Borlaug
Father of Green Revolution Iowa native and
Nobel Laureate (1970) used Biotechnology
and modern farming practices to combat world
hunger. Dr. Borlaugs development of high-yield
and disease-resistant wheat varieties bore
results in Mexico, Pakistan and India that
stretched the imagination of viable agriculture
in developing countries.  Recently, Borlaug
worked to apply farming practices and methods of
increasing food production to Asia and Africa,
and he has continued to advocate for the use
of biotechnology to combat world
famine.  Established the World Food Prize in
1987 and expanded this with the help of
John Ruan of Des Moines in 1990. Died September,
2009 at the age of 95.
3
Plant cloning
Leaf or root cutting from a plant grown into a
new plant (vegetative propagation) cloning
same genetic makeup as the donor plant.
Vegetative propagation cutting forms a mass of
non-specialized cells callus Callus
grows, divides and forms various specialized
cells (roots, stems) ?new plant.
4
Cloning approaches
Leaf cuttings Stem of a leaf in soil ?new plant,
e.g. African violets, snake plants, gloxinia, and
begonias. Some plant leaves, when cut from their
stems and placed flat on top of the soil, develop
roots at particular intervals along the cut veins
of the leaf. The parent sprouts several baby
plants, all growing vertically from the leaf at
various points along the veins, e.g.begonia and
geranium. Stem cuttings Piece of stem inserted
in soil horizontally or vertically. Roots develop
downward from stem ? new plant from top. Stem
with bud cut from plant, ? new plant from bud,
e.g begonia, gardenia, christmas cactus,
lantana, and impatiens. Budding Common in
propagation of many fruit trees. A bud, which is
an undeveloped branch, leaf or flower protruding
from the stem of the plant, cut from parent,
placed into a notch made into the parent stem,
wrapped in place and allowed to grow until it can
be removed from the parent and allowed to grow in
soil on its own. Plant division Many flowers,
such as the commonly found daylily, hostas,
certain orchids, and ferns, are cultivated
through the division of their thick roots.
Runners Wild strawberry plants, e.g., put out
horizontal stems that touch the soil. These
develop roots from which another identical
strawberry plant will grow. Grafting Cutting
from plant attached to a piece of the root, or to
the rooted stem of another plant. The pieces
become united, and grow as one plant. Desirable
properties of each plant can be mated to produce
a new hybrid plant without producing hybrid
seeds, e.g. producing dwarf fruit and oriental
trees for home landscaping
5
Meristem cloning
6
Genes and Chromosomes
This diagram shows a gene in relation to the
double helix structure of DNA and to a chromosome
(right). The chromosome is X-shaped because it is
dividing. Introns are regions often found in
eukaryote genes that are removed in the splicing
process (after the DNA is transcribed into RNA)
Only the exons encode the protein. This diagram
labels a region of only 50 or so bases as a gene.
In reality, most genes are hundreds of times
larger.
The total complement of genes in an organism or
cell is known as its genome, which may be stored
on one or more chromosomes the region of the
chromosome at which a particular gene is located
is called its locus.
7
Mendelian inheritance
Crossing between two pea plants heterozygous for
purple (B, dominant) and white (b, recessive)
blossoms
8
DNA chemistry
 
The chemical structure of a four-base fragment of
a DNA double helix
9
Mutations
DNA replication is extremely accurate, with an
error rate per site of around 1 in 10-6 to 10-10
in eukaryotes. Rare, spontaneous alterations in
the base sequence of a particular gene arise from
a number of sources, such as errors in DNA
replication and the aftermath of DNA damage.
These errors are called mutations. The cell
contains many DNA repair mechanisms for
preventing mutations and maintaining the
integrity of the genome however, in some
casessuch as breaks in both DNA strands of a
chromosome repairing physical damage to the
molecule is a higher priority than producing an
exact copy. Due to the degeneracy of the genetic
code, some mutations in protein-coding genes are
silent, or produce no change in the amino acid
sequence of the protein for which they code for
example, the codons UCU and UUC both code for
serine, so the U?C mutation has no effect on the
protein. Mutations that do have phenotypic
effects are most often neutral or deleterious to
the organism, but sometimes they confer benefits
to the organism's fitness. Mutations propagated
to the next generation lead to variations within
the population of a species. Variants of a single
gene are known as alleles, and differences in
alleles may give rise to differences in traits.
Although it is rare for the variants in a single
gene to have clearly distinguishable phenotypic
effects, certain well-defined traits are in fact
controlled by single genetic loci. A gene's most
common allele is called the wild type allele, and
rare alleles are called mutants. However, this
does not imply that the wild-type allele is the
ancestor from which the mutants are descended.
10
Biopharmaceuticals from transgenic plants

• Glycoproteins can be made (bacteria like E.
  coli cannot do this)
• Virtually unlimited amounts can be grown in the
  field rather than in expensive fermentation tanks
• There is no danger from using mammalian cells
  and tissue culture medium that might be
  contaminated with infectious agents.
• Purification is often easier

Corn is the most popular plant for these
purposes, but tobacco, tomatoes, potatoes, and
rice are also being used.
11
Biotech basics
For centuries, humankind has made improvements to
crop plants through selective breeding and
hybridization the controlled pollination of
plants.
Traditional plant breeding involves the crossing
of hundreds or thousands of genes, whereas plant
biotechnology allows for the transfer of only one
or a few desirable genes. This more precise
science allows plant breeders to develop crops
with specific beneficial traits and without
undesirable traits.
Many of these beneficial traits in new plant
varieties fight plant pests that can be
devastating. Others provide quality improvements,
such as tastier fruits and vegetables processing
advantages, such as tomatoes with higher solids
content and nutrition enhancements, such as oil
seeds that produce oils with lower saturated fat
content.
Crop improvements like these can help provide an
abundant, healthful food supply and protect our
environment for future generations.
12
Plant Biotechnology Myths Facts

• There are no biotech food products currently on
  the market.
• Biotech foods are unsafe to eat.
• Biotech foods are not regulated or tested.
• Meat, milk and eggs from livestock and poultry
  fed biotech feed products are not safe.
• Organic or conventional crops are more
  nutritious or safer than biotech crops.
• Biotech foods taste different than foods made
  from conventional crops.
• The United States does not require labeling of
  biotech foods.
• Biotech foods and crops have been rejected by
  consumers.
• The United States is the only country growing
  and consuming biotech crops.
• Biotech crops are harmful to monarch
  butterflies.
• Biotechnology is only being applied to a few
  crop varieties.
• The "pipeline" of biotech plants products is
  dried up - no new products being developed
• Biotech crops harm the environment.
• Biotech foods can't feed the world.
• Biotech crops increase food allergies.
• Using biotechnology to improve plants is not
  natural.
• Biotech companies won't disclose where field
  trials of biotech crops are being grown
• Biotech crops harm the environment.
• Biotech crops will cause "superweeds" to
  develop.

13
Animal Biotechnology Myths Facts

• Only humans, not animals can benefit from
  medical biotechnology.
• Biotech and cloned animals are still years away
  - science fiction.
• Pets do not benefit from biotechnology at all.
• Biotech and cloned animals are different from
  normal animals.
• Wild animals cannot benefit from cloning
  technology.
• Biotech will cause disease outbreaks such as
  avian flu, mad cow disease and West Nile virus.
• Organ transplants from animals are an unreal
  fantasy.
• We are just exploiting animals by applying
  biotechnology to them.
• Meat, milk and egg products from biotech animals
  are unsafe to eat.
• Biotech animals suffer more pain or distress
  than conventional animals.
• Cloned animals have higher death rates than
  conventionally-bred animals.
• Cloned animals are not as healthy as non-cloned
  animals.
• Animal cloning is not safe.
• If biotech animals or fish escape into the wild,
  they will endanger wild animals and environment.
  
• Animals are misused in research.
• The famous cloned sheep Dolly did not lead a
  full and healthy life.

14
Myth Biotech foods are unsafe to eat
Fact The Food and Drug Administration (FDA) has
determined that biotech foods and crops are as
safe as their non-biotech counterparts. The
American Medical Association, the American
Dietetic Association, and the U.S. National
Academy of Sciences have also declared biotech
foods safe for human and animal consumption. In
addition, since being introduced to U.S. markets
in 1996, not a single person or animal has become
sick from eating biotech foods. Other
international groups that have concluded biotech
foods and crops are safe are The United Nations
Food and Agriculture Organization, the World
Health Organization, the International Council
for Science, the French Food Agency, and the
British Medical Association. The European Food
Safety Authority (EFSA) has also found several
biotech varieties to be safe for human and animal
consumption.
15
Myth no biotech food products marketed
Fact 70 of processed foods on grocery store
shelves contain ingredients and oils from biotech
crops. The first biotech crop, a tomato improved
through biotechnology, was sold in 1994. The
first biotech commodity crops - an insect
resistant variety of corn - were grown and sold
in 1996. Today, the most popular biotech crops
are corn, soybean, cotton and canola.
16
C5 Plum pox resistant plums
  
Plums that have been genetically engineered to be
resistant to the plum pox virus
17
Tobacco mosaic virus resistance
Tomato plants infected with tobacco mosaic virus
(which attacks tomato plants as well as tobacco).
The plants in the back row carry an introduced
gene conferring resistance to the virus. The
resistant plants produced three times as much
fruit as the sensitive plants (front row) and the
same as control plants. (Courtesy Monsanto
Company.)
18
Pesticide resistance
Effect of the herbicide bromoxynil on tobacco
plants transformed with a bacterial gene of which
a product breaks down bromoxynil (top row) and
control plants (bottom row). "Spray blank" plants
were treated with the same spray mixture as the
others except the bromoxynil was left out.
(Courtesy of Calgene, Davis, CA.)
19
Bt Corn
Bt corn is a variant of maize, genetically
altered to express the bacterial Bt toxin,
which is poisonous to insect pests. The pest is
the European Corn Borer.
20
Bt Corn Contd

• Expressing the toxin achieved by inser-ting a
  gene from the lepidoptera pathogen Bacillus
  thuringiensis into the corn genome. This gene
  codes for a toxin causing the formation of pores
  in the larval digestive tract. These pores allow
  naturally occurring enteric bacteria such as E.
  coli and Entero-bacter to enter the hemocoel
  where they multiply and cause sepsis (Broderick
  et al., PNAS 2006). Contrary to the common notion
  that Bt toxin kills the larvae by starvation.
• Bt176 varieties were voluntarily with-drawn, in
  2001, from the list of approved varieties by the
  USEPA after finding to have little or no Bt
  expression in the ears and not found to be
  effective against second generation corn borers.
  (Current status of Bt Corn Hybrids, 2005)
• Effect on non-target invertebrates depends on
  the standard of comparison

21
Preventing Bt resistance in pests

• By law, farmers in the United States who plant
  Bt corn must plant non-Bt corn nearby. The
  non-modified fields provide a location to harbor
  pests. These refuges slow the evolution of
  resistance to the pests to the Bt pesticide.
  Doing so enables an area of the landscape where
  wild type pests will not be immediately killed.
• It is anticipated that resistance to Bt will
  evolve in the form of a recessive allele in the
  pest. Because of this, a pest that gains
  resistance will have an incredibly higher fitness
  than the wild type pest in the Bt corn fields. If
  the resistant pest is feeding in the non-Bt corn
  nearby, the resistance is neutral and offers no
  advantage to the pest over any non-resistant
  pest.
• Ensuring that there are at least some breeding
  pests nearby that are not resistant, increases
  the chance that resistant pests will choose to
  mate with a nonresistant one. Since the gene is
  recessive, all offspring will be heterozygous,
  and the offspring from that mating will not be
  resistant to Bt and therefore no longer a threat.
  Using this method scientists and farmers hope to
  keep the number of resistant genes very low, and
  utilize genetic drift to ensure that any
  resistance that does emerge does not spread.

22
The effect of Bt corn on monarchs

• A small, preliminary study done at Cornell
  University, and reported as a note in Nature in
  June 1999, indicated that monarch butterflies
  under laboratory conditions might be harmed by
  eating pollen from Bt corn plants. That
  experiment used a small number of caterpillars
  and gave them no choice about avoiding eating
  leaves that had been treated with a thick layer
  of Bt corn pollen. It did not attempt to
  duplicate real world environmental conditions.

23
The effect of Bt corn on monarchs

• There is no significant risk to monarch
  butterflies from environmental exposure to Bt
  corn, according to research conducted by a group
  of scientists coordinated by the Agricultural
  Research Service (ARS), U.S. Department of
  Agriculture. This research was published in the
  Proceedings of the National Academy of Sciences
  (PNAS).he studies in this project showed that
  monarch caterpillars have to be exposed to pollen
  levels greater than 1,000 grains/cm2 to show
  toxic effects.
• Caterpillars were found to be present on milkweed
  during the one to two weeks that pollen is shed
  by corn, but corn pollen levels on milkweed
  leaves were found to average only about 170
  pollen grains/cm2 in corn fields.
  http//www.pnas.org/cgi/content/full/211297698v1

24
Bt Cotton
Saves 3.46 M lb raw material- Conserves 1.48
Mgal fuel oil- Eliminates 2.16 M lb industrial
waste
- Transports and stores 416,000 gal insecticide
less- Conserves 604k gal fuel oil
- 1.04 million lb insecticide less 2.5 fewer
applications per acre- 416k fewer insecticide
containers- Saves 41,250 10-h workdays-
Eliminates 2,150 10-h days of aerial
application- Conserves 2.41 Mgal fuel and 93.7
Mgal water
-Accrues 168 million in economic benefits from
lower production costs and increased cotton yield
- Reduces pesticide exposure risk- Preserves
beneficial insect populations- Creates wildlife
benefits- Gives cotton producers more time for
family and community activities- Gives cotton
producers peace of mind
Produces fiber equivalent to that found in all
consumer products derived from cotton
25
Golden rice

• Progress in production of transgenic cereals for
  developing countries
• Achievement has been the introduction of genes
  that produce beta-carotene the precursor of
  vitamin A in the rice grain.
• Beta-carotene is present in the leaves of the
  rice plant, but conventional plant breeding has
  been unable to put it into the grain.
• Dr. Ingo Potrykus of the Swiss Institute of
  Plant Sciences in Zurich, with Rockefeller
  funding, transferred one bacterial and two
  daffodil genes.
• The transgenic rice grain has a light
  golden-yellow color and contains sufficient
  beta-carotene to meet human vitamin A
  requirements from rice alone.
• Potrykis has also added a gene from the French
  bean to rice that increases its iron content over
  threefold.

26
The StarLink corn controversy

• StarLink variety of Bt corn patented by Aventis
  (acquired by Bayer AG, 2002), for use in animal
  feed.
• U.S. regulatory authorities permitted StarLink
  seed but not to be grown for human consumption.
• This restriction was based on the possibility of
  an allergic reaction to the Bt protein used in
  StarLink.
• StarLink corn was subsequently found in food.
  Taco Bell taco shells was particularly well
  publicized 1
• A public relations disaster for Aventis and the
  biotechnology industry as a whole! Sales of
  StarLink seed were discontinued. Aventis
  voluntarily withdrew the registration for
  Starlink varieties in 20002
• 28 people reported apparent allergic reactions.
  However, the US Centers for Disease Control
  studied the blood of these individuals and
  concluded there was no evidence of
  hypersensitivity to Bt protein 3
• 5 weeks after the FDA/CDC declared StarLink
  safe, based on blood tests, advisers to the EPA -
  including some of the leading food allergists,
  released a thorough critique of the FDA's allergy
  test and other aspects of the StarLink
  investigation. They said the research to declare
  StarLink safe had many shortcomings including,
  lack of adequate controls, not sensitive enough
  and failed to follow standard protocols that
  helped prevent false interpretations. (FIFRA Sci.
  Advisory Panel Report 2001-09, July 01)
• Aid sent by the UN and the US to Central
  African nations also contained some StarLink
  corn. The nations involved refused to accept the
  aid.
• The southern portion of the U.S. Corn Belt
  planted the greatest amount of StarLink corn. It
  is this portion of the U.S. where corn borer
  damage creates the greatest economic loss to
  farmers.
• The US corn supply has been monitored for the
  presence of the Starlink Bt proteins since 2001.
  No positive samples have been found since 2004,
  showing that it was possible to withdraw this GM
  crop without leaving traces in the environment
  once it has been used in the field 4

27
Ethanol Contribution to Energy and Environmental
GoalsAlexander E. Farrell, Richard J. Plevin,
Brian T. Turner, Andrew D. Jones, Michael O'Hare,
Daniel M. Kammen
  SCIENCE    VOL 311 27 JANUARY 2006
www.sciencemag.org
Liquid fuel Net Fossil Inputs Net Fossil Ratio Petroleum Input GHG Emissions
MJfossil / MJfuel MJfuel produced/MJfossil MJpetroleum /MJfuel g CO2-eq. per MJfuel
Gasoline 1.19 0.84 1.10 94 
Ethanol 2005 0.774 1.30 0.04 77
CO2-intensive EtOH 0.94 1.06 0.18 91
Cellulosic ethanol 0.10 10.0 0.08 11 
http//rael.berkeley.edu/ebamm
28
Genetically modified salmon

• Armed with a gene from the ocean pout -- the
  new salmon, which originally hails from the
  Atlantic, grows twice as fast as its less endowed
  peers out at sea.
• The Food and Drug Administration has concluded
  that the new salmon is safe to eat and safe for
  the environment, suggesting that approval is
  likely at a hearing planned for later this month.
  The decision would make the fish the first
  genetically modified animal allowed for human
  consumption.

The issue has raised questions about impacts on
the environment, evolution and the future of
food.
29
Genetically modified sugar beets

• Roundup resistant
• Challenged in court September, 2010

30
Proteins produced by transgenic crops
Human growth hormone with the gene inserted into
the chloroplast DNA of tobacco plants.
Humanized antibodies against such infectious
agents as HIV respiratory syncytial virus (RSV)
sperm (a possible contraceptive) herpes simplex
virus, HSV, the cause of "cold sores" Protein
antigens to be used in vaccines An example
patient-specific antilymphoma (a cancer)
vaccines. B-cell lymphomas are clones of
malignant B cells expressing on their surface a
unique antibody molecule. Making tobacco plants
transgenic for the RNA of the variable (unique)
regions of this antibody enables them to produce
the corresponding protein. This can then be
incorporated into a vaccine in the hopes (early
trials look promising) of boosting the patient's
immune system especially the cell-mediated
branch to combat the cancer. Other useful
proteins like lysozyme and trypsin
http//images.google.com/imgres?imgurlhttp//oleg
volk.net/olegv/macro/plants/tomato.JPGimgrefurlh
ttp//kikoshouse.blogspot.com/2006/03/science-satu
rday-iv-oh-you-sweet-thing.htmlusg__8rqaLks4T2a8
o8Wi3FcOOjH7XQEh750w707sz155hlenstart8
um1tbnidGUU8gNWaI4XS2Mtbnh141tbnw133prev
/images3Fq3Dtransgenic2Bplants26hl3Den26rlz
3D1T4RNWE_enUS312US31226sa3DX26um3D1
31
Agrobacterium tumefaciens t-DNA
Invades damaged plant sites Releases transfer DNA
(t-DNA) T-DNA invades plant cells, is
incorporated into the genome Infected cells grow
rapidly to form galls The infected cells produce
food for the bacteria The invasion is DNA
transfer between kingdoms This is exploited to
introduce desirable genes This produces
transgenic plants Salt tolerant rice Golden rice
32
Transgenetics with A. tumefaciens
33
Exposure to engineered Agrobacteria
34
Reducing the rate of insect resistance development
The rate of resistance development to transgenic
plants can be reduced by either expressing
proteins with different modes of action, or
binding characteristics in one plant (gene
pyramiding) or by expressing proteins with
different modes of action or binding
characteristics in separate plants and then using
these plants either in a mosaic or sequential
fashion
35
Food Prices
Worldwide food prices have also increased
considerably. But biofuels only consume roughly
4 of the world's grains.
USDA Secretary Schafer stated last year "Higher
oil prices affect much more than just the cost of
driving they are actually one of the major
factors behind higher food costs."
The USDA and the White House Council of Economic
Advisors have stated that increased corn demand
is only responsible for "3 percent of the more
than 40 percent increase we have seen in world
food prices this year."
Ethanol is made from "dent" or yellow "feed"
corn. People accuse ethanol of increasing the
price of beer, pasta and tortillas, which is
curious because none of those products are made
from "dent" corn.
Major food companies are profiting nicely from
the food price crisis Land O'Lakes (earnings up
16), Kraft Foods (earnings up 21), Sara Lee
(net income up 55), and General Mills (profits
up 61 from 2007).
36
Galls maple trees
Galls on a maple leaf
37
Galls Rose
Rose bedeguar gall
38
Galls OAK
Oak artichoke gall caused by a wasp
39
Galls Oak Marble Galls
Cross-section (above) Oak marble galls, one with
a gall wasp exit hole and another with Phoma
gallorum fungal attack
Green stage
40
Galls Lime Nail Gall
Caused by a mite, Eriophyes tiliae, of 0.2 mm long
41
Bacterial galls
Pseudomonas syringae causes bacterial gall on
oleander and olive. ... Gall bacteria reproduce
in fissured or galled bark and are spread by ...
42
Bacterial galls rose
Agrobacterium tumefaciens manages to survive in
the rhizosphere on materials that leak from
roots. It infects wounded host plant stems and
roots.
43
Fungal galls
Fungus stem gall or "poop gall" on choke cherry
(Prunus virginiana) in Montana. The swollen stems
are caused by the fungus Dibotryon morbosum
44
Giant fungi and trees
1. The mycelium of some forest fungi can extend
enormous distances. A single individual of
Armillaria bulbosa permeates gt30 ac forest soil
in northern Michigan and may be one of the
world's largest living organisms. Some scientists
speculate that it was spawned by a single spore
thousands of years ago. Another Armillaria in
Washington was recently found to consist of a
subterranean mycelial network with erect,
above-ground mushrooms covering gt 1000 ac
2. These fungal monstrosities are rivaled in
total size and mass by a 106 acre, 6,000 ton
stand of genetically identical quaking aspen in
the Rocky Mountains. The aspen clone is
connected by a common root system, and has
literally climbed over mountains and across
meadows. Any discussion of massive clonal
colonies should also include the conjoined polyps
of coral reefs. However, the question still
remains Do these clonal colonies qualify as a
single individual, as in the 1200 ton General
Sherman tree of California's Sequoia National
Park?
45
Giant fungus oregon
Mycologists investigated if Armillaria could be
killing trees dying in the Malheur National
Forest in the Blue Mountains of eastern Oregon
and were astonished. This most recent find
was estimated to cover over 2,200 acres (890
hectares) and be at least 2,400 years old,
possibly older.