INTRODUCTION Biotechnology in agriculture has two categories: • “Improvements” to existing livestock and crops, and • Development of entirely new uses for both animals and plants (biopharming). “Improvements", include ‘input traits’ such as crops with extra resistance to insect attack, improved weed control, increase the plants tolerance to cold, drought and other environmental factor. Ex "Roundup ready" soya, "Starlite" corn, or "Frost-tolerant" tomatoes.
MOLECULAR FARMING • Molecular farming is a method used to integrate a foreign gene into plants. • Molecular farming is the term for new use plants only (not animals ) and is different in that this does not affect and has nothing to do with Food. • There are two types of Molecular Farming: Medical and non-Medical.
Medical Molecular Farming • The first synthesis of a pharmaceutically-relevant protein, human growth hormone, was described in transgenic tobacco plants in 1986. • Now, molecular farming has become commercially interesting as a method for the production of recombinant pharmaceutical proteins, in particular antibodies.
Non-Medical Molecular Farming Non-Medical Molecular Farming includes Industrial Enzymes and Polymers. • Industrial enzymes: for example laccase in transgenic maize, • Technical proteins for research purposes: for example avidin, which is also produced in maize, • Milk proteins such as human beta casein, which is produced in transgenic tomatoes, • Protein polymers: collagens, which are used for medical as well as industrial purposes.
Next… • Potentially the biggest development in this field could be the development of plants growing biodegradable plastics. • Other uses could be as Industrial oils such as hydraulic oil or highg yielding biodiesels, new solid Biofuels, new Fibres and Papers, and as agents for Bioremediation and Phytoremediation, environmentally cleaning up contamination.
WHY PLANTS? • Plants are also very flexible and can produce a wide variety of proteins. • Crop plants can synthesize a wide variety of proteins that are free of mammalian toxins and pathogens. • Crop plants produce large amounts of biomass at low cost and require limited facilities. • Crops are therefore well suited for the production of safe low-cost therapeutic proteins.
Table Comparison of Expression Systems * RT – room temperature.** N2 – culture must be maintained under nitrogen gas.
HOW IS IT DONE? • Genetic enhancement is used to introduce and express genes coding for the high value proteins.
PRODUCTION OF BRYODIN IN TOBACCO PLANT • Tobacco plants that are able to produce bryodin. • This protein, which is produced in the roots of bryonia, deactivates ribosomes and is being tested for its effect against HIV infection.
Plant Transformation • The plant leaf disc is dipped in a solution of bacteria. The bacterial "Trojan Horse" infects the edges of the leaf disc and in the process integrates the pharmaceutical protein gone into the plant genome (pict 1). • After infection the discs are placed on selection media that a flows only plant cells that carry the protein gene to survive and regenerate into plantlets. After about six weeks on selection media, a large number of plantlets that carry the pharmaceutical protein gene are visible at the edges of the original leaf disc (pict 2 & 3).
The plantlets are removed from the leaf disc and placed in clear plastic boxes that contain media that allows them to form roots (pict 4). • The rooted plantlets are placed in pots and plants are allowed to grow and produce seed. This seed can then be used for large scale production of the pharmaceutical protein (pict 5).
Protein Trafficking Following translation of the molecular ring gene, the protein will move through the endoplasmic reticulum and Golgi apparatus for processing, folding and glycosylation.
Field Production Low alkaloid plants expressing the pharmaceutical protein gone can then be produced in the field. Following harvest the tissue, the protein extracted for later use.
PRODUCTION OF SPIDER SILK PROTEINS IN PLANTS Production of transgenic plants Spider silk protein
Expression of spidroin-ELP-fusion proteins in the ER of transgenic plants
Purification of spider silk-ELP fusion proteins From transgenic plants Spidroin-ELP-fusions could be purified by addition of salt and by heat to 95% purity.