Plant Biotech Notes.pdf

- Plant genes have been manipulated for 10k years - Extensively modded by humans - Agriculture started in the mediterranean - Maize developed from teosinte through selective breeding - Took 700 years Conventional Breeding - Limited to exchanges between sam...

- Plant genes have been manipulated for 10k years - Extensively modded by humans - Agriculture started in the mediterranean - Maize developed from teosinte through selective breeding - Took 700 years Conventional Breeding - Limited to exchanges between same/related species - No guarantee of obtaining any specific gene combo from various crosses - Undesirable genes can be transferred with desirables - Long time to achieve wanted results Genetic Engineering - Allows for direct transfer of one or a few genes between closely or distant species - Can be achieved in a shorter amount of time Tools/Techniques of plant genetic engineering - Polyethylene Glycol (PEG) - Protoplasts are treated w/ PEG, inducing macro + DNA uptake by changing cell permeability - Liposomes - Foreign DNA is encapsulated in here, and protoplasts engulf the lipsomes in presence of PEG via endocytosis - Electroporation - Foreign DNA is taken up by protoplasts in a high voltage electric field - Whisker mediated - Silicon carbide fibers and protoplasts are vortexed, then the fibers make holes in the plasma membrane to allow DNA into cell - Microinjection - Foreign DNA is injected into plant protoplasts with glass needles - Laser Microbeam - UV laser makes holes in the protoplast to introduce foreign DNA - Desiccation - Dehydrated embryos can absorb foreign DNA when rehydrated with DNA solution Reporter Genes - GFP (Green Fluorescent Protein) - Cloned from jellyfish - Plant organ with GFP interacts with UV light to light up green  - GUS (E.coli’s B-glucuronidase) - Plant tissue with GUS interacts with X-Gluc + K+ferricyanide/ferrocyanide mix to turn blue - 1984 - First FDA-approved GMO tomato on the market - Polygalacturonase enhances tomato ripening, which antisense was used to suppress the expression of. - 1992-2000 - Golden Rice - Endosperm-specific promoter used to transform 3-beta carotene synthesis genes - Could help population with vitamin A deficiency Video Notes (To Be Organized) - Plant Biotechnology Applications: Medicine, disease/pest resistance, drought tolerance, biofuel, nutrition, high yield, comparative genomics, phytosensor(detecting toxins)/phytoremediation(removing chemicals from soil) - Tobacco plants were genetically modified to prod. “Plantibodies” that inactivated Ebola: “pharmed” for pharmaceuticals - Every gene must have a promoter, unless its an operon - Promoter -> GUS -> Terminator - GUS -> mRNA -> Protein - Reporters (GUS, GFP, LUC) are not selectable markers - Gene Gun/agrobacterium transformation = most popular methods to transfer foreign genes to plants - Gene Gun - high velocity particle delivery system - Helium travels(600 psi) -> rupture disk -> macrocarrier(has gold-covered DNA) -> stopping screen(stops macrocarrier from destroying particles) -> particles travel down to target shelf -> DNA integrates into host organism’s chromosomes - Agrobacterium transformation - TI (tumor inducing) plasmid contains virulence genes, makes sure at least one sequence is excised -> TDNA travels through channel to infect plant nucleus -> integrated into gene - Grown gall disease - tumor-like mass in plants - Disarmed plasmid - plasmid that no longer contains disease gene - Delivery system -> integration -> recovery - PCR is necessary to confirm integration -