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Tissue Culture
Did you ever have a plant that was so
unique or so beautiful that you wished
you had hundreds or thousands of them
to enjoy or to sell? Plant tissue culture
(micropropagation) is a technique which
will do just that for us. We are going
to discuss this tool which is used so
extensively in the nursery business and
in plant biotechnology. It is a fascinating
and useful tool which allows the rapid
production of many genetically identical
plants using relatively small amounts
of space, supplies and time.
Basically the technique consists of taking
a piece of a plant (such as a stem tip,
node, meristem, embryo, or even a seed)
and placing it in a sterile, (usually
gel-based) nutrient medium where it multiplies.
The formulation of the growth medium is
changed depending upon whether you are
trying to get the plant to produce undifferentiated
callus tissue, multiply the number of
plantlets, grow roots, or multiply embryos
for "artificial seed".
For many who
become superficially aware of the technique
it seems shrouded in mystery and is shrugged
off as too technical to be of concern.
Actually, it is no more of a mystery than
taking a cutting of your favorite house
plant and growing it to share with a friend.
As for being technical, you can begin
plant tissue culture with as little as
a cookbook approach and a feeling for
sterile technique.
Some people have visions of scientists
doing plant tissue cultures in white gowns
and masks in hospital-clean environments.
Such conditions are excessive. While it
is true that mold spores, bacteria, and
other contaminants will grow and overrun
a culture, air that is not moving has
a minimum of contaminants. In addition,
disinfection of implements, work surface
and nearby areas helps eliminate contaminants.
The guidelines for preparation
and the laboratory protocol provided here
are given as a place to begin. Included
with is a limited discussion of some of
the many options you have as you explore
micropropagation.
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We can discuss these in more depth if you
have questions, concerns or related experiences
to share. I would be particularly interested
in success and challenges you may have had
or are currently having in your classroom.
Some suggestions are given for the following
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Selecting
plant sources. Some species, or even clones
are easier to grow in culture than others.
Some respond reluctantly to culture, some
do not respond at all, and many plants
have never been tried.
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Choosing a growth
medium (price, convenience, type of plant
and purpose of the micropropagation all
enter into this decision.) How important
are the kinds of hormones used? On limited
scale, media ingredients are available
at the grocery and health food stores. |
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Suggestions for media
preparation and sterilization. There are
alternatives to sterilization in a pressure
cooker or an autoclave. |
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Methods for cleaning,
storing and manipulating explants (plant
pieces to be cultured). |
Given certain basics there
are many options for procedure, equipment and
supplies for plant tissue culture. Some of your
decisions will be based upon the amount of time,
money and space you have. Other decisions will
be based upon why you are doing plant tissue
culture and what you expect as a result (more
plants?) . Catalogs, such as Sigma, Carolina
Biological, or Edmund Scientific are good reference
and they are for purchasing needed materials..
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Equipment Used in Tissue Culture
Laboratory
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Analytical balance (for
weighing nutrients for media) |
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Graduated cylinders and pipettes (for
measuring stock solutions) |
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pH meter (to regulate pH of media) |
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Hot plate or stove (to heat and dissolve
gelling agent) |
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Glass containers (for heating and dissolving
media) |
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Dispensing devices (to dispense equal
quantities of media) |
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A Still or de-ionizer (water needed
for media) |
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Pressure steam sterilizer (for sterilizing
instruments and media) |
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Transfer instruments (forceps, scalpels
spatulas, blades) |
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Refrigerator (storage of chemicals
and stock solutions) |
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Stereo-microscope (use for meristem
culture) |
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Laminar Flow Hood (provide a sterile
area for transfers during initiation and sub-culturing) |
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| Adantages
of Propagation by Tissue Culture |
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The
elimination of diseases and the production
of disease free plantlets |
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The
rapid production of large numbers
of genetically identical plantlets |
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Introduction
of new varieties and or genotypes |
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Preservation
of germplasm |
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Production
of haploid plants which can be used
for plant breeding |
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Production
of plantlets from species in which
plant development from seed is difficult
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Requirements for Plant Tissue Culture |
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a
washing area (vessels and planting
material are cleaned, plantlets may
be weaned) |
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a media preparation room
( preparation of media, storage and
sterilization) |
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a aseptic transfer area
(initiation and sub-culturing of plantlets) |
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an incubator or a culture
room (provide plantlets in culture with
temperature and light
requirement) |
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