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February 5, 2001


Farmers Confident in Biotech; Step Backward with Canadian



Feb. 5/01 From a press release (Via Agnet)

With first-hand knowledge of the value and benefits biotechnology delivers
to their farms and to consumers around the globe, U.S. farmers are again
poised to plant a large share of their corn, cotton and soybean acres with
biotech seed in 2001.

A report from 13 of the nation's leading agricultural and commodity
organizations and the Council for Biotechnology Information (CBI)
indicates the recent controversy that has swirled around one particular
product -- StarLink corn -- has done little to dampen farmers' enthusiasm
for biotechnology. "Farmers across the country know the benefits of
biotechnology are real and very significant -- not just for agriculture
but for consumers, as well," notes Lee Klein, a farmer from Battle Creek,
Neb., and president of the National Corn Growers Association. "For
example, corn and cotton products that carry the Bt trait provide these
crops with natural resistance to pests that can cause tremendous damage,"
he adds. "This in-plant protection provides a terrific environmental
benefit because it lets farmers use less pesticide in a more precise
manner. Herbicide resistance in certain corn hybrids, and in some
varieties of cotton and soybeans, gives farmers the opportunity to
effectively control weeds with limited applications of crop protection
products. In addition to this important benefit, these crops help farmers
deliver a healthier supply of food and fiber to consumers around the
world. These are just a few of the reasons why American farmers strongly
support continued access to products of biotechnology." Ron Heck, who
grows corn and soybeans outside the central-Iowa community of Perry is
just one of those farmers. "The publicity surrounding StarLink hasn't
shaken our confidence in the value of biotechnology one bit," he states.
"Our farm is located very close to the western Corn Belt, where corn
borers can be a real problem," Heck adds. "Bt corn provides a safe,
economical and environmentally friendly option for controlling these
pests. That's why we plan to maximize our use of Bt hybrids approved for
food, feed and export for 2001."

A significant percentage of U.S. corn growers seem to agree with Heck. In
an online survey by AgWeb.com, conducted the week of November 17, 2000,
farmers were asked how the recent controversy surrounding biotechnology
would affect their seed corn selections for 2001. Results showed that a
total of 45 percent would plant either the same or a greater percentage of
biotech corn for the coming year. In comparison, 29 percent said they
would plant a reduced percentage or no biotech corn in 2001. The use of
biotech products in soybeans has increased steadily over the past few
years. It appears that trend will continue in 2001. "Our biotech products
are an important part of our business," says David Thompson, director of
marketing communications with Stine Seed in Adel, Iowa. "Our sales grew
last year and we're anticipating they'll grow again this year. Farmers
have been very pleased with products of biotechnology and that certainly
has been reflected in our sales figures." Scott Beck, vice president of
Beck's Hybrids in Atlanta, Ind., says sales of Roundup Ready soybean seed
are up 8 percent over last year. Roundup Ready varieties, he adds, account
for 94.5 percent of the company's soybean seed sales. The adoption of
biotech products in the cotton industry over the past few years has been
rapid and widespread. Annual USDA Cotton Varieties Planted Reports show
plantings of biotech varieties have increased from 13 percent of total
cotton acreage in 1996 to 70 percent in 2000. At least one seed company
expects this trend to continue. "Looking ahead to the 2001 growing season,
we're again seeing increased sales of biotech varieties," notes Steve M.
Hawkins, president of Delta and Pine Land Company. "We have several
options for farmers to choose from -- offering insect protection,
herbicide tolerance and the combination of traits -- and farmers are
finding these products offer significant value." Donna Winters is one of
those farmers who have seen the value of biotechnology on her Lake
Providence, La. farm.

"Biotechnology has proven to be a vital tool," she says. "Products like
Bt cotton allow us to apply fewer pesticides to the crop. This reduced
chemical use and the fact that we now make a lot fewer trips over our
fields with equipment are big pluses for the environment. These products
also improve our efficiency tremendously. Economically and
environmentally, I don't think either American farmers or consumers can
afford to lose access to the products of biotechnology." Farmers would be
the first to point out that they obviously don't operate in a vacuum.
That's why they are heartened by research that indicates consumers have
not changed their food consumption behavior in response to the StarLink
corn incident.

In a major study conducted this past fall by the Grocery Manufacturers
Association and Dr. Thomas Hoban of North Carolina State University,
consumers were asked the following question: "During the past few months,
have you done anything or taken any action because of any concerns you may
have about genetically modified foods?" The overwhelming majority -- 95
percent -- responded, "No."

"As farmers, we always have to be aware first and foremost of any
concerns consumers might have about our nation's supply of food and
fiber," says Tony Anderson, a Mt. Sterling, Ohio, soybean farmer who is
also the current president of the American Soybean Association. "But as we
work to help increase consumers' awareness of biotechnology, we're
confident they'll understand what we've known for several years:
Biotechnology is an advancement in science that helps us produce safer,
more nutritious foods and higher quality fibers while enhancing our
stewardship of our nation's critical land and water resources."


The February 2001 issue of ISB News Report at http://www.isb.vt.edu
has following stories:

USDA Announces Risk Assessment Research Grants
Cloned Gaur Dies
Emerging Technologies In Plant Biotechnology
Engineered Chloroplasts Snip Out Antibiotic Resistance Genes
The Vision Of An Edible Vaccine For Hepatitis B Starts To Come Into Focus
Plants That Detect Landmines, And Other Biosensors
Transgenic Milk Containing Lysostaphin: A Possible Cure For Mastitis?
Premarket Notice Concerning Bioengineered Foods
Identity Preservation And Product Segregation Procedures


Terence Corcoran February 6, 2001 Editoria, National Post (Via Agnet)

Columnist Corcoran says that the long, fruitful march of scientific
progress took a step backward in Ottawa yesterday with the report of The
Royal Society of Canada on the future of Canadian food biotechnology. You
could tell there was trouble right from the opening pages. The first
referenced source in the 265-page report, the first "scientific" footnote
in a work packed with scientific pretense and commissioned by the
government of Canada, cites Cloning the Buddha: The Moral Impact of

This is no accident or coincidence. Cloning the Buddha is a mystical
anti-science track by New Age spiritualist Richard Heinberg. Corcoran says
that Maude Barlow probably sleeps with a copy at her bedside, right next
to Jeremy Rifkin's anti-biotech work, The Biotech Century. A typical
Heinberg statement is the following false moral quagmire: "The fear is
that we will fail in our attempts [at a biotech revolution] and trigger a
universal biological catastrophe; or that, even if we succeed, in doing so
we will erode and destroy the human soul, and perhaps the very soul of
nature. Either way, the implications are enormous."

Corcoran says that the reference to Cloning the Buddha is inexplicable and
indefensible in a so-called expert panel report of leading scientists
commissioned by three departments of the federal government. Unless, of
course, the original intent was to turn the science upside down; then it's
perfectly clear why Mr. Heinberg is the first authority in the report.
Titled Elements of Precaution, the Royal Society's report is a universal
catalogue of regulatory overkill, a virtual Joy of Regulating textbook
that in the end seeks to drown biotechnology, genetically modified (GM)
food and scientific progress in a bathtub of regulation.

The list of scientific tripwires proposed by the society -- if they were
ever implemented -- would bog regulators, industry and science down for
decades. Everything would need to be tested, scientists would have to
prove everything is safe, which inevitably means proving the negative,
proving that biotech is not a hazard to human health and biodiversity. A
typical recommendation is the following: "The primary burden of proof is
upon those who would deploy these food biotechnology products to carry out
the full range of tests necessary to demonstrate reliably that they do not
pose unacceptable risks."

Greenpeace and Ms. Barlow's Council of Canadians immediately hailed the
report, pleased by suggestions that the government undertake "exhaustive,
long-term testing for ecological effects of biotechnology products." That
prescription alone implies stalling GM progress while regulators send out
for more and more studies to prove there are no risks. Indeed, the
society's blue-print reads like a plot to kill off biotechnology. Cause of
death: the precautionary principle.

Corcoran says that from the mysticism of Heinberg in the opening chapter,
the report takes 200 pages -- packed with regulatory ideas -- before it
gets down to its hard-core premise. The precautionary principle is the
environmental movement's Magna Carta, its charter of rights and freedom to
regulate and prevent any action if there is some perceived threat to
health, safety and bio-diversity. If there's a risk that something will go
wrong, regulators don't need to be scientifically certain that it will go
wrong. Under the precautionary principle, scientific progress must stop
unless it can be proven to be safe.

The Royal Society panel created its own definitions of the precautionary
principle. A sample: "If there are scientific data (even though
incomplete, contested, or preliminary) -- plausible scientific hypotheses
or models (even though contested) -- together with significant levels of
uncertainty, that establish a reasonable prima facie case for the
possibility of serious harm (with respect to reversibility, remediation,
spatial and temporal scale, complexity and connectivity), then
precautionary action is justified."

That's bad enough. The panel then went on to say that even if the
scientific community concludes that there is no risk, regulators should
still plow ahead. Their example: the world-wide scare over mad cow
disease. "Even when the available scientific evidence fails to establish a
risk as nothing other than 'remote,' where there is a prima facie case of
serious risk, significant (in this case highly costly) precautionary
action is warranted." The precautionary principle, in various forms and
definitions, is now embedded in most Canadian and international law. It
claims to be a risk management tool. In fact, it distorts and conceals
risks. As one writer put it, the precautionary principle is a "rhetorical
weapon." It's purpose is to "exaggerate the risks associated with economic
endeavor and conceal the risks arising from the exercise of political

The Royal Society report embodies the full perversity of the precautionary
principle. Through 265 pages it posits and imagines hundreds of risks,
urges delay and research, and not once does it even mention the risks
associated with the regulations it proposes. What are the risks of not
developing GM foods and other biotech products? How many lives will be
lost if the world does not get better, cheaper, safer food?

Canada's biotech rules will eventually be set by the government, based on
advice from responsible councils. This is not one of them.

Fighting Diseases With Bioengineered Foods?

By PHIL LEMPERT Los Angeles Times Monday, February 5, 2001

Susan Harlander sees a future in which we will choose what to eat based on
our own genetic makeup. With the benefit of genetic testing, we would know
whether we carried genes that predisposed us to illnesses such as cancer,
heart disease, Alzheimer's and diabetes. We'd then eat foods--many of them
the products of genetic engineering--that would be designed to help
prevent or cure those diseases. That future may not be that far off, says
Harlander, a consultant in New Brighton, Minn. Already, many foods
produced with the help of genetic engineering are on supermarket shelves.
Today, more than 55% of all soybeans and nearly half of all corn produced
in the United States is genetically modified for insect resistance, lower
herbicide use or greater yield. And gene-splicing techniques have been
used to improve a variety of foods, such as beer and tomatoes.

Most of the controversy surrounding bioengineered crops and food has
focused on such issues as labeling and the risk of potential environmental
and health problems. Biotechnology is the process in which a specific
gene--or blueprint of a trait--is isolated and removed from one organism,
then relocated into the DNA of another organism to replicate that trait.
Last May, the U.S. Food and Drug Administration issued new rules that
require a mandatory review of all foods that are produced through genetic
engineering. While the debate over bioengineered foods continues, the food
industry is working to stock supermarkets with these genetically
engineered foods that will claim to enrich, maintain and prolong your life.

Here's a look at some of the new bioengineered foods that will be arriving
in the supermarkets in the next two to three years: Golden rice is a
genetically modified rice with increased levels of beta-carotene and other
carotenoids, which has been created to help fight vitamin A deficiency, a
leading cause of blindness. Scientists at Alabama A&M University at
Normal, Ala., are working to remove the protein that is the source of
peanut allergies. Being able to alter the saturation levels of oil means
healthier peanut, canola, soybean and sunflower oils; and according to
Harlander, such products are awaiting approval by the FDA. New soybean
oils, for example, have been developed that are more than 80%
monounsaturated and contain 33% less saturated fat than olive oil. Such
oils will be helpful to people seeking to reduce their cholesterol levels.
Lycopene, found mostly in tomatoes, is highly regarded as one of the most
effective antioxidants. Harvard researchers discovered in 1995 that
lycopene lowered the risk of prostate cancer. Now, researchers at the
University of London/Royal Holloway Hospital have created tomatoes that
contain three times the amount of this cancer-fighting, heart-protecting

And researchers have isolated the specific DNA trait that produces
caffeine in young tea leaves, which will allow them to develop plants that
are naturally deficient in caffeine. The plants would retain the same
flavor and aroma as their caffeine counterparts.
* * * Phil Lempert hosts a national syndicated radio show and is the
food correspondent for NBC's "Today" show. He can be reached at
PLempert@aol.com. Copyright 2000 Los Angeles Times

Bioengineering for the mouths of babes
Missy Globerman Redherring.com, January 19, 2001


Deep beneath a five-floor labyrinth of laboratories on the fringe of
Cornell University's campus lurks a plant biologist who means to save
children from the doctor's needle. "See that stuff inside," says Charles
Arntzen, as he shows us a triple-stuffed British cookie, not unlike an
Oreo, that he'd purchased the other day in London's Heathrow Airport. "In
just a few years, my friend, that will be the dried concentrate powder of
hepatitis B vaccine made from tomatoes, mushed into a tasty crème. A great
snack for kids everywhere, wouldn't you say?"

Dr. Arntzen, 59, may be as popular with the Nobel Prize Committee as he
will be with the kids if his bioengineered fruits and vegetables can ward
off disease in countries too poor to refrigerate and store conventional
vaccines. Granted, such vaccines are the core of preventative medicine,
having already rid the world of smallpox and come close to ridding it of
polio. Yet even now, one infant in five goes unvaccinated against a slew
of other plagues, including diphtheria, tetanus, and measles. How
different matters might be if you could load a canoe with vaccine-carrying
bananas, potatoes, or tomatoes and paddle it down the Amazon or Congo

PRESCRIPTION FOOD Perhaps if Monsanto (NYSE: MON) had come out with such
fruits of mercy before unveiling crops that make their own pesticides, the
company would have disarmed critics of bioengineered food before they
could build a head of steam. "It's very hard to be pro-infant mortality,"
Dr. Arntzen notes dryly.

Dr. Arntzen is alive to the problem of public perception. Having grown up
on a farm in Minnesota, he still remembers his dairy-farmer neighbors
fighting an earlier wonder food, margarine, hyping its supposed health
hazards and getting Washington to pass laws to discourage its sale. Dr.
Arntzen cringes at the memory, saying that the ingrained horror of
technology has already slowed the development of vaccine technology. His
first financial backer, Britain's Axis Genetics, went bankrupt last fall,
in part because of political opposition to so-called Frankenfood.

Dr. Arntzen, a grandfatherly man with a bit of a paunch, did his
undergraduate work at the University of Minnesota. His first love was
rocks, and it was only after taking a break from college to marry his
wife, Kathy, that he switched to molecular biology, graduating in 1965. He
went on to earn a doctorate in cell physiology from Purdue University,
then spent five years at the University of Illinois, punctuated by
visiting professorships overseas. He became a globe-trotter, beginning in
India, where he served in a U.S. government mission from 1976 to 1981 to
teach villagers how to grow drought-resistant crops.

His eureka moment came in 1992, at a floating market just north of
Bangkok. As Dr. Arntzen watched, a mother swabbed the lips of her crying
baby with a bit of banana, and the baby stopped crying. "I knew at that
point it would be totally logical to put something useful into that
fruit," Dr. Arntzen remembers. "It's obvious children like it, and it's a
sterile little syringe that every developing country can grow."

Dr. Arntzen got the insight because his mind was prepared. He had spent
four years in the mid-'80s working in DuPont (NYSE: DD)'s program for
bioengineered crops, assembling a medley of skills that won him an unusual
double appointment: dean of the school of agriculture at Texas A&M
University and adjunct professor of physiology at the University of Texas
Medical School.

To prove that he could engineer a plant to make a foreign protein, Dr.
Arntzen began with a banana's genome and added DNA coding for an enzyme
that turns blue in a particular reagent. He placed it into a seed, planted
the seed, harvested the fruit, and put a piece of it into the reagent. The
result was a blue banana. His next move was to make a banana manufacture
an antigen -- a protein on the surface of a germ that the immune system
can recognize and attack.

The work "involves a lot of patience," Dr. Arntzen notes. He pulls a petri
dish off a cart, picks up a baby banana plant taking root in it, and shows
it to me. It took his lab two and a half years to make that seedling,
which produces an antigen from the hepatitis B virus. It will be eight
more years before he gets a ripe banana. (To get more bananas of his
earlier design is easy -- just take cuttings from the existing
10-foot-high plants.)

Besides needing to prove that a plant could make antigens on command, Dr.
Arntzen also had to overturn the conventional wisdom that an oral vaccine
requires a whole virus, not just one protein isolated from the virus.
Trials in rodents cast doubt on this prejudice. Then, in 1997, Dr. Arntzen
published his first human trial, involving a potato-borne antigen from
strains of Escherichia coli that often cause food poisoning. All 11
volunteers who ate the potato developed an immune response to the bacteria.

PRIMING THE PUMP Dr. Arntzen has also broken new ground in understanding
how the gut accepts vaccination. The process, involving what doctors call
mucosal immunity, offers a great advantage: the proteins generate
antibodies that hide in the body's mucous membranes, serving as the first
line of defense against invaders. Although it works wonderfully in animal
trials, researchers still need to show that it works perfectly in humans.
Above all, doctors must be reassured that the vaccine will never be
recognized merely as food, inducing tolerance for the antigen, and thus
the germ.

Edible vaccines may well be safer than injectable ones because their
manufacture has absolutely nothing to do with the germs that cause
disease. "Remember when kids used to get sick from the flu shot? Aha! No
more," he exults. "The proteins we isolate cannot actually cause the
diseases" they're supposed to protect against. The first federal
regulations for plant-based vaccines are now being hammered out, with Dr.
Arntzen's help, and he hopes to get U.S. approval for two vaccines --
against hepatitis B and the Norwalk virus, cause of so-called traveler's
diarrhea -- within two years. "It may really be more like five years," he
says, "but I would never send this vaccine technology out to other
countries without approval here first."

Dennis Lang, head of the National Institutes of Health's Institute of
Allergy and Infectious Diseases, has known Dr. Arntzen since their days at
DuPont. "A real innovator and a really smart guy, and someday when people
are talking about edible vaccines around the dinner table, his name will
surely be mentioned as one of the first pioneers of this technology," Dr.
Lang says. "But the potential of this produce is still anybody's guess,
and remember that developing countries won't just grow these drugs in the
woods. They will have to be licensed, regulated, controlled, and purchased
from the Americans with the patent."

So far the main technical problem has been assuring the dosage. Raw,
unprocessed fruits and vegetables cannot deliver consistent amounts of
vaccine, so the plan now is to puree, can, or dry the banana vaccines into
chips. The tomatoes will be dried and reconstituted into sauce. The first
vats of processed tomatoes can be seen already on Cornell's campus in its
Food Science Department.

Dr. Arntzen is leaving Cornell's Boyce Thompson Institute for Plant
Research, where he has been president, for yet another academic perch, at
Arizona State University, in Tempe. There he will collaborate with Dow
AgroSciences on the development of edible vaccines for livestock. As he
and his wife plan their move to the bottom of a desert cliff in Arizona,
he makes a point of learning to enjoy life outside the lab, in year-round
sun. He gave his skis to the Salvation Army and bought a BMW Z3
convertible; now he buffs his golf game and listens daily to the song
"Don't Worry, Be Happy."

"I think every so often about why I got into this line of work. I always
remember first and foremost that I love food and working with food. Then I
think of the dangers of the work." He ruminates for a moment. "The most
dangerous thing we as scientists can now do about vaccines is to do
nothing. The world cannot afford to let this development opportunity pass
us by."


From Cot Curves to Genomics. How Gene Cloning Established New Concepts in
Plant Biology

Robert B. Goldberg*, Department of Molecular, Cell, and Developmental
Biology, University of California, Los Angeles, California 90095-1606
Plant Physiol, January 2001, Vol. 125, pp. 4-8

It is difficult to imagine carrying out plant research without personal
computers, the Internet, GenBank, e-mail, cell phones, gene cloning,
microchips, whole genomic sequences, expressed sequence tags, RFLPs, PCR,
knock-outs, Arabidopsis, reverse genetics, transgenic plants, and
molecular biology "kits" that are ready-made to carry out almost any type
of DNA manipulation experiment imaginable.

The plant world in 1975 was vastly different from the one in which we, as
plant scientists, operate in today. The International Society of Plant
Molecular Biologists did not exist. International forums such as the Plant
Molecular Biology Gordon Conference, the Plant-Oriented Keystone Symposia,
and the Plant Molecular Biology Congress had not been established. One of
the largest gatherings of plant scientists occurred at the annual meetings
of the American Society of Plant Physiologists and seldom more than 20 or
30 scientists attended the nucleic acids section in which the most
exciting plant molecular biology results were presented. The "real world"
was different as well. The Vietnam War had just ended, the Cold War with
the Soviet Union raged on, the Berlin Wall split Europe into East and
West, and the world economy was in an inflationary spiral due to the
emergence of the oil cartel that sent the prices of gasoline skyrocketing.

Genetic engineering had been "invented" by Stanley Cohen and Herbert Boyer
2 years earlier (11) and was still limited to an elite number of labs that
understood bacterial genetics, had the plasmid vectors for DNA cloning,
and had access to the enzymes that we purchase in cloning "kits" today.
Procedures for cDNA cloning, creating libraries of large eukaryotic
genomes, and isolating structural genes had not yet been published.
Genetic engineering was as controversial then as genetically modified
organisms are today. The Asilomar Conference took place in 1975, and
scientists who wanted to use the emerging tools of genetic engineering
were required to follow strict, self-imposed guidelines that specified the
conditions under which DNA manipulations could be carried out in the
laboratory. Demonstrations occurred across the globe forecasting that
"monsters" would be created by the new gene splicing techniques and one
city (Cambridge, MA) attempted to ban genetic engineering altogether.
Nevertheless, it was a magical time to be studying basic plant processes.
For the first time, there was a dream that one could finally "see" a plant
gene and begin to unravel the complexity of plant processes at the genome

DOWN IN THE PRECLONING ERA Plant genomes were investigated in the mid- to
late-1970s by quantitative DNA reassociation tools (i.e. Cot curves) that
had their origins in the 1960s when the principles of DNA denaturation and
renaturation were pioneered at the Carnegie Institution of Washington by
Roy Britten and his associates (7, 8)principles that are still used today
each time a gel blot or microchip experiment is carried out, a primer Tm
is calculated, or PCR conditions are punched into a thermocycler. Plant
genomes had been shown to contain repetitive DNA sequences in the
mid-1960s and were, therefore, considered to be "eukaryotic-like" and
similar to animal genomes in that respect (7, 8). In 1975, genome
organization was the "code word" for those of us who studied "genomics"
and it was determined that plant genomes had many families of repetitive
sequences and that these repeats varied in copy number and arrangement in
the genome (17, 20). These repeats were shown to be both scattered around
the genome and localized in long clusters and they were also shown to be
flanked by complex single-copy sequences (17, 20). Neither these repeats
nor any flanking single-copy DNA had been cloned or sequenced at this
time. In fact, DNA sequencing procedures (29, 33) had not yet been
invented and plant DNA sequences had not yet been cloned (3). However, the
general concepts of plant genome organization that were derived from DNA
reassociation studies have stood the test of time and have been
illuminated in great detail by a knowledge of the actual DNA sequences
that span each Arabidopsis chromosome (5).

During this same period, important principles of plant gene activity were
being established in global terms by the use of RNA-excess/DNA-RNA
hybridization techniques (i.e. Rot curves) with either cDNA or genomic
single-copy DNA probes (21, 22, 24, 25). The technique of subtraction
hybridization (or cascade hybridization as it was first referred to in the
literature) was established by Bill Timberlake in this era using
kinetically fractionated cDNA populations (36). Both cDNA and genomic
single-copy DNA subtraction procedures were used by many of us to
investigate developmental changes in plant mRNA populations (21, 22, 24,
25). Several important concepts emerged about higher plant cells in this
precloning population hybridization era. First, it became clear that
plants contained a complex set of nuclear RNAs and that only about 25% of
this complexity was represented in the corresponding mRNA population (21).
Today, we know that the additional complexity in the nuclear RNA
represents primarily unprocessed introns in primary transcripts. However,
this was not understood at the time because plant genes had not yet been
cloned and sequenced, and introns had not yet been discovered in any
eukaryotic gene. Second, it became clear that a large number of genes were
active in plant cells and that these genes were highly regulated in the
plant life cycle (21, 24, 25). Each plant organ system was shown to have a
unique set of active genes and it was estimated that approximately 60,000
genes were required to program and maintain the entire life cycle of the
tobacco plant (24). This estimate of the number of tobacco genes has stood
the test of time for plants with large genomes (i.e. corn) and,
considering the "bluntness" of the tools used and assumptions that had to
be made (e.g. average mRNA size), is not that far off from the 25,000
genes that has been shown by sequencing to be present in the small
Arabidopsis genome (5). Finally, it was established that mRNA populations
contained sequences with varying degrees of prevalence and that both
transcriptional and posttranscriptional processes established the mRNA
sequence sets present in various plant organs and tissue types. By the end
of the Cot and Rot curve era (mid-1980s), it was clear that plant cells
resembled animal cells with respect to the number of genes and the
complexity of gene regulatory processes. It was not known, however, how
any individual gene was regulated or how sets of genes were co-expressed
in space and time.

PLANT GENES CAN BE CLONED! By the end of the 1970s, exciting new
procedures were developed by Tom Maniatis and others to construct cDNA
clones of specific eukaryotic mRNAs and isolate the corresponding genes
from the genome (26, 27). In addition, techniques were devised to sequence
DNA segments (29, 33), visualize genes directly in the electron microscope
in association with their RNAs (i.e. R loops; 38), and detect specific DNA
fragments and mRNAs using DNA and RNA gel blots, respectively (1, 34).
These procedures established a new revolution in molecular biology
because, for the first time, the structures of individual genes could be
studied and their expression patterns, mechanisms of regulation, and
evolutionary origins analyzed. This was an exciting period and the most
surprising and startling observation made with the new DNA cloning
techniques was that the coding regions of eukaryotic genes were
interrupted by non-coding sequences (23)! New words, intron and exon, were
introduced into the molecular biology lexicon (19) and posttranscriptional
splicing mechanisms were hypothesized and studied (23). Only a few plant
scientists at that time had any experience with bacterial genetics, the
new recombinant DNA techniques, or access to enzymes required for DNA
cloning and manipulation. In fact, most of us did not know a restriction
enzyme from a ligase and had to learn from "scratch" how to streak and
grow bacterial cells in order to attempt to clone plant DNA sequences! In
the 1970s and 1980s (as well as today) plant scientists were playing
"catch-up" with their animal counterparts and were competing for a meager
pot of money. It was during this time that Joe Key played a huge role in
establishing the U.S. Department of Agriculture Competitive Research
Grants Program after many years of fighting the U.S. Department of
Agriculture bureaucracy and Congress. This Program has made a major impact
over the past 25 years in keeping plant sciences in the forefront of
pioneering research. Rumors began to circulate in the late 1970s that
plant DNA could not be cloned. One well-known plant molecular biologist
(who will remain anonymous) went from meeting to meeting like Paul Revere
declaring that plant DNA was "different" from animal or bacterial DNA and
that it could not be cloned! John Bedbrook and colleagues in Dick
Flavell's lab in Cambridge, England soon showed that this was not the case
and demonstrated directly that plant DNA could be cloned and replicated in
bacteria just like the DNA from other organisms (3). They reported their
results in 1979 at a meeting in Minneapolis and the era of plant gene
cloning began with the successful cloning of ribosomal DNA and telomeric
repeated sequences from wheat (3). A pioneering principle was
establishedplant DNA was similar to that of all other organisms and could
be manipulated using the same enzymes, cells, and vector systems.

Soon thereafter, libraries of many plant genomes were constructed and, in
the early 1980s, were made available to plant scientists around the world
(16, 35). In addition, the first plant structural genes were cloned,
sequenced, and visualized in the electron microscope (16, 35). These
genes, encoding seed storage proteins (16, 35) and the small subunit of
ribulose bisphosphate carboxylase (4), were shown to contain introns
similar to those in animal genes, which supported the notion that plant
cells had genetic processes similar to those in animals. It was also
demonstrated that plant genes were located relatively close to each other
on plant chromosomes (approximately every 4-6 kb) and that genes with
different expression patterns were interspersed among each other, implying
that each functioned as an independent unit (16)a suggestion that was
verified during the post-transformation era (9, 31, 32).

During the same period, cDNA libraries were constructed for almost every
imaginable plant organ system and developmental state, and cDNA clones
representing prevalent plant mRNAs, such as those encoding seed proteins,
light-regulated proteins, hormone-induced proteins, and cell wall proteins
were identified. These cDNA clones were used to demonstrate directly that
both transcriptional and posttranscriptional processes played a role in
controlling plant gene expression, but that the primary control for most
plant genes was at the level of transcription. In addition, the vast array
of cDNAs that became available and were sequenced and studied in the 1980s
began to illuminate a range of plant developmental, metabolic, and
biochemical processes. The age of understanding "how to make a plant" had

PICKING APART PLANT GENES As the new era of plant gene cloning began,
another revolution was occurring in several labs that were engaged in a
fierce competitive battle to be the first to transform plant cells. The
laboratories of Jeff Schell and Marc Van Montagu (Gent, Belgium), Rob
Schilperoort (Leiden, The Netherlands), Mary-Dell Chilton and Michael
Bevan (Washington University, St. Louis; Cambridge University, UK), and
Rob Fraley, Steve Rogers, and Rob Horsch (Monsanto, St. Louis) were
utilizing the new recombinant DNA techniques to construct Agrobacterium
tumafaciens T-DNA vectors that could be used to introduce new genes into
plant cells. In the mid-1970s, Mary-Dell Chilton had shown that A.
tumafaciens T-DNA was integrated into the chromosomes of plant cells (10),
setting the stage for the revolution in plant genetic engineering that
continues to this day.

In 1983, the Gent, Monsanto, and Washington/Cambridge groups showed
independently that T-DNA vectors could be used to transfer bacterial
antibiotic resistance genes into plant cells and that these genes could be
expressed if engineered with the correct promoters (6, 12, 18). Much to
the surprise of everyone in the plant research world, a different group,
headed by Tim Hall, demonstrated that the phaseolin seed storage protein
gene from french beans could be transferred to sunflower cells and be
expressed (31). This now-famous (or infamous) "sunbean" plant made the
front page of the New York Times and was proclaimed in Time to be a
"glowing achievement... the first time a gene from one plant had been
inserted into the chromosomes of an unrelated species and made to express
itself." The sunbean experiment was reported initially at the first
University of California (Los Angeles) Keystone Meeting on Plant Molecular
Biology that I organized in April of 1983 and was greeted at the time by a
now-famous plant cell biologist (who I will not name) as "nonsense!"
Nevertheless, it showed for the first time that gene cloning and A.
tumafaciens transformation techniques could be combined to transfer
foreign genes into plant cells and study their function. The age of plant
genetic engineering and gene manipulation had begun!

FROM PHENOTYPE TO GENE Throughout the 1980s and 1990s, many plant genes
were cloned and investigated in transformed cells in order to understand
the mechanisms regulating their expression. Numerous plant promoters were
characterized and DNA sequence elements programming transcription in
specific developmental states were uncovered. The prediction of earlier
experiments on the structure and organization of plant genes proved
correct and a major new concept emergedplant genes functioned as
independent units and contained regulatory regions that could program
their correct expression in foreign cell environments. These experiments
set the stage for engineering new crops with novel traits that are
produced at specific times during the plant life cycle (28). A major
switch in plant gene cloning occurred in the beginning of the late 1980s
and early 1990s. Many interesting genes that produced novel phenotypes
were being uncovered in corn and Arabidopsis using genetic approaches that
were being adopted rapidly by plant scientists. Because their products
were unknown and/or very rare, it was not possible to use conventional
cloning methods to isolate these genes. Several pioneering procedures were
invented that circumvented this problem and enabled a wide range of plant
genes to be cloned. First, T-DNA was shown to act as a mutagen in plant
cells and, as such, could be used as a tag to identify and clone genes
that specified novel phenotypes (15). Ken Feldmann and his colleagues
established a novel seed transformation method to obtain large numbers of
T-DNA transformed Arabidopsis lines and this method was used to identify
important plant genes, such as those involved in the control of floral
organ identity and hormone perception (14, 15). In my opinion, this was
one of the most important advances in plant biology in the past 25 years
because it allowed, for the first time, a relatively simple way to clone
plant genes associated with fascinating mutant phenotypes. The
availability of Ken Feldmann's T-DNA lines caused numerous investigators
(including myself) to adopt Arabidopsis as a model system and opened up
many new problems in plant biology to investigation. It also paved the way
to the reverse genetics approaches in use todayidentifying mutant lines
associated with randomly sequenced genes (30).

A second approach to cloning plant genes was also being developed at the
same time. During the 1980s, Nina Fedoroff and Sue Wessler cloned the corn
Ac and Ds transposable elements (13). This pioneering experiment paved the
way for using transposons to tag and capture novel plant genes for which
only a phenotype could be identified. The transposon tagging and gene
cloning procedure complemented the T-DNA approach and led to the
identification of many important new genes in several plants including
corn, snapdragon, and Arabidopsis (37). It also became possible in the
1990s to use map-based cloning strategies to identify and clone plant
genesparticularly in Arabidopsis because of its small genome size (2).
With the completion of the Arabidopsis Genome Project last fall, and the
identification of 30,000 single nucleotide polymorphisms in the
Arabidopsis genome, map-based cloning of plant genes should permit the
identification of any gene for which there is a mutant phenotypeeven those
induced by chemical mutagens such as ethyl methane sulfonate.

BACK TO THE FUTURE Looking back, in 1975 plant molecular biologists were
asking questions about the number of genes in plant chromosomes and how
these genes are regulated in development. We were using precloning tools
of DNA and RNA hybridization that gave precise answers, but which could
not focus in on specific genes. The questions addressed then are being
addressed once again today in the genomics age. In a sense, we have come
full circle in trying to understand how plant chromosomes are constructed
and how populations of genes are expressed in various cells, tissues, and
organs. We progressed from studying populations of genes and mRNAs to
investigating individual cloned genes and mRNAs to using high throughput
experiments with arrays of thousands of specific genes in order to uncover
the secrets of plant cells. Twenty years after the cloning of the first
plant DNA segments (3), the genomes of Arabidopsis and rice have been
sequenced and numerous expressed sequence tag sequencing projects have
uncovered tens of thousands of mRNAs in a wide range of plants (5). It is
remarkable that the era of gene cloning is coming to an end. Nevertheless,
the challenges are no less daunting and are even more complex: What are
the functions of all plant genes and how is the information in plant
genomes utilized in order to program plant development from fertilization
to seed dormancy?

* E-mail bobg@ucla.edu; fax 310-825-8201.

LITERATURE CITED (cut here...see