Biotechnology in Agriculture Chapter 11

Learning Outcomes 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

Asexual Plant Propagation 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 Arabidopsis thaliana – an herbaceous plant, related to radishes, that serves as a model organism for many plant genetic engineering studies Crossbreeding – pollination between plants of different phenotypes, or varieties 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. Runners – long, vine-like stems that grow along the soil’s surface 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

11.1 Review Questions Which of the following are examples of asexual plant propagation: PTC, selective breeding, stem cuttings, leaf cuttings, or runner? How is length or width added to a plant? 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 Plant growth regulators – another name for plant hormones Auxin – a plant hormone produced primarily in shoot tips that regulates cell elongation and leaf development Cytokinin – a class of hormones that regulates plant cell division Explants – sections or pieces of a plant that are grown in or on sterile plant tissue culture media Ethylene – a plant hormone that regulates fruit ripening and leaf development Abscisic acid – a plant hormone that regulates bud development and seed dormancy Phytochrome – a pigment that acts like a hormone to control flowering

11.2 Review Questions What is another name for plant hormones? Auxin is responsible for what kind of plant growth regulation? Cytokinin is responsible for what kind of plant growth regulation? 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 Agriculture – the practice of growing and harvesting animal or plant crops for food, fuel, fibers, or other useful products Horticulture – the practice of growing plants for ornamental purposes Inbreeding – the breeding of closely related organisms 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 Hydroponics – the practice of growing plants in a soil-less, water-based medium Plant-based pharmaceutical (PBP) – a human pharmaceutical produced in plants; also called plant-made pharmaceutical (PMP)

11.3 Review Questions What is it called when very closely related animals are bred? Why is it discouraged? Name two advantages of growing plants hydroponically. 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 Genomic DNA (gDNA) – the chromosomal DNA of a cell Agrobacterium tumefaciens (A. tumefaciens) – a bacterium that transfers the “Ti plasmid” to certain plant species, resulting in a plant disease called crown gall; used in plant genetic engineering 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

11.4 Review Questions Which is larger, gDNA or pDNA, and by how much? Plant DNA is difficult to get out of plant cells. List a few “tricks” used by technicians to isolate plant DNA. Why is the bacteruim, A. tumefaciens, of interest to biotechnologists? 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 Transgenic plants – plants that contain genes from another species; also called genetically engineered or genetically modified plants 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 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 What is the name of the naturally occurring bacterium and the plasmid that can infect plants and transfer DNA molecules? Name at least two selection genes that are used to confirm that Ti plasmid transformation has occurred. How does GUS act as a selection gene? Why are so many plant genetic-engineering experiments conducted with Aradibopsis, even though it has little, if any, economic value?

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