Genetic Engineering (Can be fun!)

1. Genetic Engineering(Can be fun!) Basic Bio 100 M. Marshall Spring 2014

2. The Genetic engineering story . . . . . like that seen in other branches of science is a story of serendipity combined with some clever thinking. Techniques were developed that allowed, among others (not necessarily in chronological or importance order): The sequencing of genomic DNA. The isolation and manipulation of specific DNA sequences (genes). The ability to isolate and clone isolated “foreign” genes in bacteria and select out those clones containing the specific DNA sequences of interest. The ability to isolate plant cells and grow them in culture as unspecialized “callus” tissue and in some cases protoplast cultures. The ability to insert specific genes into plant cells and have their proteins expressed by their new host cell. The ability to regrow these “transformed” cultures into whole plants

3. Sequencing speed has increased by leaps and bounds . . And many many others. The amount of DNA sequence information available today is increasing at a rate that necessitated the founding of a whole new area of information management: Bioinformatics

4. GE bacteria and plants are possible because scientists were able to combine discoveries from different areas of biology, some of which originally seemed to have no obvious practical application. Bacterial plasmids, small circles of extra-chromosomal DNA, were discovered in experiments exploring bacterial drug resistance. These DNAs contain genes, that can produce proteins, just like those in chromosomes. They are also easy to remove, manipulate, and then re-insert into their bacterial hosts. Someone realized they could be used as”vectors” to move foreign genes into bacteria which would then be able to replicate them and produce the foreign gene products as if they were their own.

5. Bacterial “restriction” enzymes were discovered that cut DNA internally, but only at particular recognition sites. Some create a staggered cut that always leaves the same “sticky ends”making it easy to join different DNAs cut with the same enzyme together. These became a tool that “gene jockeys” use to isolate genes and place them into plasmids for cloning.

6. Extra Photo 12.16x

7. Figure 12.2

8. The DNA donor cell can be animal, plant, or whatever. The process shown here allows a desired human protein to be produced by engineered bacteria so that it can be purified and used as a medicine. The process is A LOT more complex than shown here, but this shows the main events

9. This eliminates problems with insulin resistance that often developed over time as the recipient’s immune system developed an immune response to the bovine and porcine insulin used previously. Today type I (insulin-dependent) diabetics take human insulin, “Humulin” produced by special strains of transformed E.coli.

10. Attempts have been made, so far without much success, to cure human genetic defects by using viruses as vectors to replace faulty genes with normal copies.

11. Plant Genetic Engineering - Is misunderstood by many which has lead to much misinformation being circulated about the risks and benefits involved.

12. Plants have been selectively modified using artificial selection (cross breeding, hybridizing, and selection) since agriculture began. Genetic engineering (GE) expands this process to include gene transfer from other species, even across Kingdoms. Plants are modified to resist pests, grow better, or produce crops that are of higher quality than before. Shown below “Golden rice”plants produce β-carotene (provitamin A) not only in green tissues (the leaves) but also in the seed endosperm. Vitamin A deficiency causes childhood blindness in societies that consume high quantities of normal rice.

13. GE plants have been created to produce crops with a greater shelf life (tomatoes), be resistant to herbicides (cotton), and to create their own insecticides. This last objective has been achieved in several plant species, field corn and cotton to name two. Corn earworm (a moth larva) is one insect controlled in this manner.

14. Bacillus thuringensis (Bt) Bt is a bacillus that produces a spore when conditions are unfavorable for growth. Sporulation is accompanied by the production of a protein crystal outside of the spore. The cell degrades leaving only the dormant spores and crystals behind. The bacterium is specifically toxic to lepidopterans (butterfly and moth larvae) and some coleopterans as well. The ingested protein crystals (Cry protein toxin) perforates the larvae gut, killing it and allowing the spores to germinate and use the larvae for food, and the cycle starts over when the new bacteria are done growing throughout the larval carcass. The bacteria were sold as a biological “pesticide” dust for years and still are.

15. Cry toxin activity

16. The Cry toxin will kill larvae when ingested regardless of the source • Bt corn and cotton contain the toxin gene and their cells make their own toxin constitutively (all the time). The bacteria “middle man” has been eliminated. • This was only possible because a method was developed to transfer the toxin gene into plant cells.

17. Plant tumors, or crown galls, were found to be produced by a bacterium that transfers a plasmid (the Ti plasmid) to the host plant cells. Plasmid genes cause the cells so transformed to grow out of control; as they do so, they produce several odd amino acids that only Agrobacterium can use as food. The last piece of the plant GE puzzle is provided by Agrobacterium tumefaciens, the Crown Gall bacterium. Modified Ti plasmids with the disease-producing genes removed can be used as vectors to move desirable foreign genes, such as that for the BtCry toxin, into plants.

18. Plant tissue culture provides the cells to be transformed Plant pieces are grown into unspecialized “callus” cultures which are transformed with the Agrobacterium Ti plasmid containing the Cry toxin gene and the callus cells are then induced to reform into plantlets. The process selects for the survival of only those plant cells that have taken up the Ti plasmid and are expressing the proteins its genes code for. These are then induced to reform into plants with the proper sequence of hormone treatments.

19. If the foreign gene codes for the insect toxin from Bacillus thuringiensis, the plants would now be able to make their own protein insecticide (Bt toxin).

20. Franken - food! Unfortunately there is much misinformation and paranoia surrounding the production and consumption of GMO crops.

21. GMO Phobia Genetically engineered organisms are considered ”unnatural” and therefore potentially harmful by some individuals and organizations. These fears are especially acute when it comes to attitudes toward modified food crops. The main fears are: • Overt toxicity • Carcinogenicity • Allergenicity • Environmental damage The USDA, FDA and EPA all have a role in regulating and approving GM food organisms for release into the environment and for human and animal feed. They are some of the most highly regulated materials in our society, yet concerns and paranoia continue.

22. This paranoia would be funny if the consequences weren’t so serious. The potential benefits of this technology are especially promising for less prosperous third world countries where the high-input agriculture we practice here is not practicable. But many countries have drunk the cool aid of anti-GMO paranoia to the point where they refuse to let GMO crops be grown or sold there. GMO crops have been highly tested for toxicity and allergenicity to the point where newly developed varieties are considered safe by definition, which mean less testing these days. That alarms the paranoids even more. Many are categorically convinced that GMOs are dangerous and any data to the contrary is dismissed as industry propaganda.

23. So now in addition to food . . . we export fear to the rest of the world

24. Some sample craziness