Transcript Plant tissue culture

Biotechnology in
Agriculture
Chapter 11
Learning Outcomes
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Describe the role of meristematic tissue in propagating plants by
various asexual methods
Outline and discuss the process of plant tissue culture, including the
importance of the different hormones involved, and identify the
advantages and challenges of plant tissue culture
Give specific examples of agricultural and horticultural biotechnology
applications, including genetically modified organism (GMO) crops,
hydroponics, and plant-made pharmaceuticals
Explain how genomic and plasmid DNA can be isolated from cells,
including the additional steps required for plant cells
Summarize the methods used to produce transgenic plants and
explain the selection processes for identifying transformed plant cells
11.1 Cloning Plants
Using breeding techniques, a plant biotechnologist can produce variety
in the offspring of selected parental plants.
Asexual Plant Propagation
Identical offspring are produced by a single parent.
Methods of Asexual Plant Propagation
Vocabulary
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Arabidopsis thaliana – an herbaceous plant, related to radishes, that serves
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Crossbreeding – pollination between plants of different phenotypes, or
varieties
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Asexual plant propagation – a process by which identical offspring are
produced by a single parent; methods include the cutting of leaves and stems,
and plant tissue culture, etc.
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Runners – long, vine-like stems that grow along the soil’s surface
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Plant tissue culture (PTC) – the process of growing small pieces of plants
into small plantlets in or on sterile plant tissue culture medium; plant tissue
culture has all of the required nutrients, chemicals, and hormones to promote
cell division and specialization
as a model organism for many plant genetic engineering studies
11.1 Review Questions
1.
Which of the following are examples of asexual plant propagation:
PTC, selective breeding, stem cuttings, leaf cuttings, or runner?
2.
How is length or width added to a plant?
3.
Leaf or stem cuttings must include a least some of what kind of
tissue to form new roots?
11.2 Cloning by Plant Tissue Culture
In PTC a few cells are grown in sterile media, with sugar,
vitamins, and correct hormones.
Hormone Function in Plants
Plant Growth Regulators
Auxin
Cytokinin
Starting a Tissue Culture
Hormones, agar, nutrients
Advantages of Plant Tissue Culture Propagation
More plantlets are produced.
Produces clones of the parent with no variations.
Some plants do not propagate will other ways.
Factors to Consider in Plant Tissue Culture Propagation
The species and variety of plant material
The medium and medium ingredients
The preparation of plant samples, medium, and equipment (sterility and
temperatures, etc.)
Vocabulary
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Plant growth regulators – another name for plant hormones
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Auxin – a plant hormone produced primarily in shoot tips that regulates cell
elongation and leaf development
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Cytokinin – a class of hormones that regulates plant cell division
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Explants – sections or pieces of a plant that are grown in or on sterile plant tissue
culture media
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Ethylene – a plant hormone that regulates fruit ripening and leaf development
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Abscisic acid – a plant hormone that regulates bud development and seed dormancy
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Phytochrome – a pigment that acts like a hormone to control flowering
11.2 Review Questions
1.
What is another name for plant hormones?
2.
Auxin is responsible for what kind of plant growth regulation?
Cytokinin is responsible for what kind of plant growth regulation?
3.
How can a plant tissue culturist know that an explant is beginning
to respond to the hormones in the PTC media?
11.3 New Applications of Biotech in Agriculture and Horticulture
Selective breeding of livestock and plant crops has been practiced for
centuries.
New techniques are now being applied.
Genetic Testing and Gene Transfer
With DNA fingerprinting, breeders can test parent animals and plants for
several beneficial genes and recognize several undesirable genes.
Benefits of Selective Breeding and Propagation
• Animals:
• Improved nutritional value
• Fewer feed additives
• Increased growth rate
• Plants:
• Resistant to selected viruses
• Higher nutritional content
• Less fertilizer or herbicide
• Less environmental impact from run-off pollution
Hydroponics: An Alternative Plant Growing Method
Soil-less, water-based medium in which to grow plants
Plant-Made (Plant-Based) Pharmaceuticals
Since the 1970s, human proteins have been made in bacterial,
fungal, or mammalian cell cultures.
Vocabulary
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Agriculture – the practice of growing and harvesting animal or plant crops for food, fuel,
fibers, or other useful products
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Horticulture – the practice of growing plants for ornamental purposes
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Inbreeding – the breeding of closely related organisms
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Bacillus thuringiensis (B. thuringienses or BT) – the bacterium from which the Bt gene
was originally isolated; the Bt gene codes for the production of a compound that is toxic to
insects
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Hydroponics – the practice of growing plants in a soil-less, water-based medium
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Plant-based pharmaceutical (PBP) – a human pharmaceutical produced in plants; also
called plant-made pharmaceutical (PMP)
11.3 Review Questions
1.
What is it called when very closely related animals are bred?
Why is it discouraged?
2.
Name two advantages of growing plants hydroponically.
3.
How are PMPs related to genetically engineered organisms?
11.4 Isolating DNA from Plant Cells
The DNA used in plant biotechnology applications may be
genomic DNA (gDNA) or plasmid DNA (pDNA).
Isolating Genomic DNA
Cells must be burst open.
Proteins are precipitated and removed from solution.
RNA is destroyed.
Remaining DNA is precipitated.
Isolating Plasmid DNA
Plasmid isolation kits
Buffers for plasmid isolation
Vocabulary
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Genomic DNA (gDNA) – the chromosomal DNA of a cell
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Agrobacterium tumefaciens (A. tumefaciens) – a bacterium that transfers the “Ti
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Ti plasmid – a plasmid found in Agrobacterium tumefaciens that is used to carry genes
into plants, with the goal that the recipient plants will gain new phenotypes
plasmid” to certain plant species, resulting in a plant disease called crown gall; used in
plant genetic engineering
11.4 Review Questions
1.
Which is larger, gDNA or pDNA, and by how much?
2.
Plant DNA is difficult to get out of plant cells. List a few “tricks” used
by technicians to isolate plant DNA.
3.
Why is the bacteruim, A. tumefaciens, of interest to biotechnologists?
4.
Why is Ti plasmid of interest to biotechnicians?
11.5 Plant Genetic Engineering
Using A. tumefaciens to
Genetically Engineer Plants
Transforming Agrobacterium.
Before A. tumefaciens can be used
to transform a plant, its Ti plasmid
must be transformed with the
gene(s) of interest.
Ti Plasmid. The Ti plasmid has two selection genes on it, NPT II and beta-D-glucuronidase (GUS), so that when it
gets into plant cells, the plasmid transfer can be recognized. Cells receiving this plasmid will be able to survive on
kanamycin-containing agar (from NPT II expression). They will also be able to convert a white carbohydrate in the
medium to a blue color (due to GUS expression), which makes the entire colony blue, allowing the researcher to
ascertain successful DNA transfer.
Arabidopsis thaliana, a Model Organism for Plant Genetic Engineering
Arabidopsis thaliana has been the target of plant genetic engineering
studies.
Vocabulary
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Transgenic plants – plants that contain genes from another species; also called
genetically engineered or genetically modified plants
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NPT II (neomycin phosphotransferase) gene – a gene that codes for the
production of the enzyme, neomycin phosphotransferase, which gives a cell
resistance to the antibiotic kanamycin
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GUS gene – a gene that codes for an enzyme called beta-glucuronidase, an
enzyme that breaks down the carbohydrate, X-Gluc, into a blue product
11.5 Review Questions
1.
What is the name of the naturally occurring bacterium and the
plasmid that can infect plants and transfer DNA molecules?
2.
Name at least two selection genes that are used to confirm that
Ti plasmid transformation has occurred.
3.
How does GUS act as a selection gene?
4.
Why are so many plant genetic-engineering experiments
conducted with Aradibopsis, even though it has little, if any,
economic value?
Questions and Comments?