Finding New Ways To Protect and Preserve
Plant Genetic Resources
Cryopreserved garlic shoot tips. The tips were excised
from plants growing in tissue culture, immersed in droplets of cryoprotecting
solution, and frozen in liquid nitrogen to preserve genetic diversity
of the specimens.
"To protect and to serve" is the motto for the Los Angeles Police
Department. The motto for the National Center for Genetic Resources Preservation
in Fort Collins, Colorado, could be "to protect and to preserve."
The ARS center stores more than 450,000
samples of plant and animal germplasm, such as seed and semen. The Plant Germplasm
Preservation Research Unit (PGPRU) at the center researches ways to manage the
collection more efficiently and to get germplasm to survive longer.
Making a collection of germplasm is called genebankingand it's not as
simple as just filing seeds away. Some germplasm does not store well, so scientists
must discover ways to extend shelf life. To reduce the cost of genebanking,
scientists must ensure that the collection doesn't include unnecessary germplasm
but is still large enough to include valuable genes. The PGPRU researchers are
using genetic tools and mathematical models to help "right-size" the
collection so that it includes much of what researchers and plant breeders need
but is not too unwieldy.
The following examples show how PGPRU scientists solve genebanking problems
to provide breeders, researchers, and growers the genetic resources needed for
healthy agricultural systems at minimum cost to taxpayers.
Preserving the Unpreservable
These wild-rice plants are being grown in the greenhouse
in a study of how their seeds develop. Plant physiologist Christina Walters
(left) and technician Lisa Hill collect the seeds and label flowers.
Consumers love wild rice. But breeders have had their work cut out for them
to bring this crop into mainstream cultivation,because the seeds don't survive
"The seeds are called 'recalcitrant' because they can't be preserved easily.
Breeders are losing valuable genetic stocks every year," according to plant
physiologist Christina Walters, research leader of the unit.
Usually wild-rice breeders store seeds in a conventional refrigerator. Unfortunately,
seeds stored this way may not survive even a full year and thus need to be grown
out every spring.
Walters and her staff are trying to find better methods. They have found that
the water content of seeds can be optimizedmaking them neither too wet
nor too dry. Some drying slows down seed aging and germination but does not
hurt the seed. Drying also means less chance that lethal freezing will occur,
so the seeds can survive at lower temperatures for longer times. Walters's group
has shown that wild-rice seeds can be stored for at least 3 years at 5°C.
Storing wild-rice seeds longer is possible, but it is labor intensive, because
the embryo must be dissected from the seed and then cryopreserved, that is,
rapidly cooled in liquid nitrogen.
In studies of genetic diversity of bristlecone pines,
molecular markers allow unique specimens or populations of trees to be
identified. Here, geneticist Chris Richards loads an automated DNA sequencer
for genetic analysis.
Which Garlic Is Which?
When germplasm like wild-rice seed is labor-intensive to preserve, it makes
sense to store only what is needed and not waste efforts by inadvertently storing
the same thing over and over again. But for some plants, like garlic, preventing
duplication is a challenge.
Because garlic does not reproduce by seeds, varieties are kept as clonesthat
is, individuals with exactly the same genes (like identical twins). This means
that bulbs from California Early garlic, for example, are clones of the same
individual, whether they're purchased in Washington State or Washington, D.C.
Different varieties of garlic arose from mutations that occurred over the years.
Growers gave names to the new garlic types, but over time the names changed
or similar names were given to different garlics. It is now impossible to distinguish
among garlics by variety name. It is also difficult to identify garlics by appearance,
since cloves from the same bulb grow differently in different locations.
These garlic shoot tips were immersed in droplets of cryoprotecting
solution, then plunged into liquid nitrogen to preserve genetic diversity
of the vegetatively propagated germplasm.
To reconcile the problem, a team led by plant physiologist Gayle M. Volk used
DNA markers to examine groups of garlic varieties and determine how much diversity
exists among them. The team conducted a genetic analysis of 211 garlic accessions
using what's known as "amplified fragment length polymorphisms." They
found that many accessions bearing different names were in fact virtually indistinguishable.
But there were also plenty of unique accessions. The garlics can now be tracked
by genetic identity rather than name, helping genebankers to preserve the most
diverse garlics first.
Bristlecone pines could be considered Earth's oldest living inhabitants, since
some of them may be 5,000 years old. Unfortunately, the bristlecone pine is
being threatened by white pine blister rust, which has devastated populations
of white pines in the Northwest and is rapidly moving through the Rockies. Currently,
germplasm of this legacy species is not preserved in any genebank.
Geneticist Christopher M. Richards, in collaboration with U.S. Forest Service
scientist Anna Schoettle, is trying to assess diversity of bristlecone pines
and to identify, for collection, germplasm that represents what is in the wild.
Using a cryomicroscope, plant physiologist Gayle Volk
(left) and technician Ann Caspersen examine changes that occur in cell
structure during cryoprotection.
"We have to identify individual trees that have important genes, such
as resistance to blister rust," explains Richards. "Pollen from this
species travels large distances, so there's a lot of genetic mixing. We need
to look at genetic diversity tree by tree."
In his work detecting genetic diversity, Richards is using DNA-based markers
called "retrotransposons," or "jumping genes." Sequences
of these genes help uncover hidden variation within species.
Retrotransposons are genetic elements in the genomes of many living organisms,
including humans, that are replicated in response to some unknown environmental
cue. After several replications, there is an increasingly long trail of DNA
insertions, which can be used as a DNA fingerprint. This new DNA sequencing
tool is helping to distinguish tree families and identify individual trees with
important genetic differences.
Technician Remi Bonnart (foreground) flash-freezes plant
shoot tips while plant physiologist Christina Walters places cryopreserved
materials into cryovats for long-term storage. Droplets of cryoprotecting
solution hold shoot tips onto foil strips. The strips are plunged into
liquid nitrogen slush that is formed by placing liquid nitrogen in a vacuum.
Just like bristlecone pines, grapes are threatened by a pathogen, in this case
Xylella fastidiosa. This bacterium, which causes deadly Pierce's disease,
is spread by the glassy-winged sharpshooter, an insect that has invaded grape-growing
regions around Davis, California, and elsewhere. Davis is also home to one of
two ARS regional genebanks that specialize in preserving grapes. (The Fort Collins
center acts as a backup to all the regional genebanks.)
Says plant physiologist Leigh E. Towill, "There are more than 3,300 lines
of grapes preserved by ARS, and all are field- maintained," making them
vulnerable to disease or other natural disasters. Towill's main focus is cryopreserving
plant cuttings from the field collections.
"Cryopreserving these lines is an effective backup strategy that will
save money and ensure the germplasm is safe."
To buy time while other scientists battle Pierce's disease, Towill has developed
a method to store grape scions for 18 months at 3°C. That way, if
something were to happen to the Davis field collection, the grape lines could
be quickly restored by simply rooting vines that were in storage.By David Elstein, Agricultural
Research Service Information Staff.
Plant physiologist Gayle Volk inspects garlic plants growing
in culture. Shoot tips of garlics were cryopreserved because these plants
represent the greatest genetic diversity in the collection.
This research is part of Plant, Microbial, and Insect Genetic Resources,
Genomics, and Genetic Improvement, an ARS National Program (#301) described
on the World Wide Web at www.nps.ars.usda.gov
Christina Walters, Gayle
M. Volk, Christopher M. Richards,
and Leigh E. Towill are in
the USDA-ARS Plant Germplasm Preservation Research Unit, National
Center for Genetic Resources Preservation, 1111 South Mason St., Fort Collins,
CO 80521-4500; phone (970) 495-3200, fax (970) 221-1427.
"Finding New Ways To Protect and Preserve Plant Genetic Resources" was
published in the April 2005 issue of Agricultural