Biotechnology & SynBio

1. Biotechnology & SynBio By C. Kohn, Waterford WI

2. Better Bushels • Imagine if we had a crop that produced its own pesticide… • We wouldn’t have to spray fields with a chemical pesticide • This would save farmers money and protect the environment • It would also reduce the pathogens that affect our crops, reducing the need for other protective treatments while making fields more productive. • Would this be a good thing or a bad thing? Discuss

3. Bt Corn • Such a product does exist and has been used for over 15 years. • In fact, 45% of the corn grown in the US is genetically modified. • The most famous example of this is Bt Corn. • Left: Control • Right: Bt Corn

4. Bt Corn • Bt Corn is a GMO, or Genetically Modified Organism. • GMO: a plant or animal that has been genetically modified through the addition of a small amount of genetic material from other organisms. • In a GMO, genetic material from another organism is inserted into the plant or animal genome. • These genes can come from any living source, including bacteria, fungi, and other organisms.

5. Bt = Bacillus thuringiensis • In the case of Bt Corn, an inserted gene for a natural insecticide came from Bacillus thuringiensis, a bacterium found naturally in the soil. • B. thuringiensis bacteria naturally produce a toxin (the Bt delta toxin) which kills specific predatory insects during the larval stage. • It does not harm other insects in the way broad-spectrum insecticides do, making it an ideal replacement to synthetic chemical pesticides. • Bt was actually available as a separate pesticide since 1960 and has an excellent safety record, making it an ideal choice as a GMO.

6. Production of Bt Corn • Production of Bt Corn was relatively straightforward: • The gene for the Bt toxin was sequenced and identified. • The gene was removed from the B. thuringiensis genome using a restriction enzyme. • The genome of corn was spliced using the same restriction enzyme. • The gene was inserted and made permanent using DNA ligase. • The modified corn genome was inserted into a corn cell nucleus. • The corn cell, when it divided, produced the Bt gene along with the rest of the corn’s genome.

7. Bt in action. • Because Bt corn has the gene for the Bt toxin, it produces this protein just like any other protein in a corn cell. • When an insect ingests the Bt toxin protein produced by the corn, the Bt toxin binds to the stomach wall of the insect. • Within hours the stomach wall is broken down by the toxin.

8. Bt & Monarchs • Concern has been raised about the impact of Bt corn on monarch butterflies. • Research by the USDA’s Agricultural Research Service has shown that other than an early version of Bt Corn (which has since been replaced), the impact on monarchs is negligible and insignificant. • Plus, the alternative to Bt corn is the use of chemical pesticides, which are far more harmful to butterflies.

9. Is It Safe? • Bt corn was approved by the USDA for human consumption in 1995. Is it safe? • This might be a good question, given the Bt toxin kills insects by destroying their intestinal tracts • “Delta endotoxins and VIPs produced by the currently available events all are rapidly broken down in the stomach and thus are not potential food allergens.” – Colorado State University • i.e. your own stomach will rapidly break down the toxins before they can affect you • Bt corn is considered generally safe a not a threat to consumers. • It is regulated by both the EPA and FDA for human and environmental safety. • It has been used for over 15 years with no record of serious issue.

10. Biotechnology • Bt Corn has become the poster child of biotechnologies made possible by recombinant DNA. • Recombinant DNA has made the science of biotechnology possible. • Recombinant DNA: when genes from two different species are combined and introduced into a cell • Biotechnology: the manipulation of the genetics of organisms to make useful products • Biotechnology is not necessarily a new science • Selective breeding of livestock and the use of microbes to make wine are ancient examples of biotechnology • However, with the use of recombinant DNA and other technologies, biotechnology has changed modern life.

11. Making Recombinant DNA • Production of recombinant DNA is similar regardless of what you are producing. • First, a gene must be cut using a restriction enzyme (a chemical scissors for DNA that always cuts at the same sequence of bases) • Copies of DNA always yield the same restriction fragments when exposed to a restriction enzyme (meaning DNA copies are always cut in a predictable way).

12. Making Recombinant DNA • If a restriction enzyme cuts DNA in such a way that a single-stranded portion remains, this is called a “sticky end” • Sticky ends are important because they allow the addition of new genes so long as they have the complementary sequence to the sticky ends • E.g. a new gene would have to have a TTAA sticky end to ‘fit’ inside these restriction fragments.

13. Creation of Recombinant DNA • 1. A restriction enzyme cuts DNA • 2. Restriction fragments are created • 3. A new gene with complementary sticky ends is inserted. • 4. DNA ligase (an enzyme) permanently seals the new gene into the genome. RestrictionEnzyme DNALigase

14. DNA Ligase • DNA ligase enzyme is necessary to “cement” the new gene into the genome. • Without DNA ligase, the bond is only temporary. • DNA Ligase is the “super glue” that makes a bond permanent • Once DNA ligase has formed a permanent bond with the new gene and the original genome (the “vector), we have recombinant DNA. • A cloning vector is the DNA that carries the inserted gene DNALigase

15. Biofuels • Scientists are currently working to develop GM organisms that can convert cellulose (in plant cell walls) into (such as ethanol and biodiesel). • Scientists are working to identify the genes responsible for more efficient breakdown of cellulose and how and where to insert these genes into the genome of modified organisms like E. coli and yeast. • The hope is that an organism could be developed to either breakdown cellulose more efficiently than we currently can with yeast produced through selective breeding.

16. Synthetic Biology • The use of recombinant DNA in biotechnology has led to the rise of a new science: synthetic biology. • Synthetic Biology: a science combining engineering, biotechnology, and biology in order to create new biological organisms that do not exist naturally in the environment. • In a sense, synthetic biology is the combination of genes from different sources to create a “super organism” that can do all the things we need it to do. • By selectively choosing and inserting genes, the hope is that we can create organisms from scratch that can address the most pressing needs facing society.

17. Examples of Syn-Bio Attempts • Synthetic Biology has been used so far to address the following: • The production of more productive microbes for the rumens of cattle • Production of more potent and effective vaccines • Production of bio-engineered drugs that target and destroy cancer tumors • Bioremediation microbes that can quickly clean up an oil spill or toxic waste • Biofuel-producing microbes that can convert plant feedstocks into more useful substances (cellulose into glucose for ethanol or oil for biodiesel).

18. The Moral and Ethical Implications • If a product of synthetic biology were to escape into earth’s fragile, complex and highly interdependent ecosystem, it could have devastating effects on all naturally occurring organisms. • Our “super organisms” could become super-invasive, seriously disrupting natural processes. • The unintentional release of synthetically engineered organisms could have damaging side effects. • Many who oppose this type of research fear that this science could be abused may fall into the hands of terrorist groups and rogue nations.

19. Current Research • Watch the following video • Recombinant DNA, as advanced as it is, is even being phased out by advances in SynBio. • Use of recombinant DNA was sort of like writing a book using words and sentences from other books. • Scientists now are trying to create genes from scratch that could be inserted into cells, enabling them to write their “book” from scratch rather than trying to find the genes responsible for the traits they are trying to create. .

20. SynBio & Malaria • One of the first major successes with Synthetic Biology was the production of precursor to the compound artemisinin. • This compound has been shown to be effective against strains of malaria that are resistant to more widely-used drugs • However, this drug was far too expensive to produce • "By inserting genes from three separate organisms into the E. coli, we're creating a bacterial strain that can produce the artemisininprecursor." Jay Keasling, lead researcher on the project.

21. SynBio Malaria Medicine • Keasling and his team took genes from yeast and from the sweet wormwood tree (the source of the drug compound) and inserted them into E. coli. • As a result, the yield of the artemisinin precursor in that strain of E. coli was increased by 10,000 times compared tothat of the wormwood tree. • Right: Keasling and the wormwood tree • Photo courtesy of: www.lbl.gov

22. Biofuel of the Future • If we were to create a similar organism as the malaria-drug E. coli bacterium, what traits would it need? • Think/Pair/Share – what kinds of genes would we need to insert into yeast or E. coli in order to produce the ideal biofuel microbe? • What aspects of this work might interfere with the function of this modified organism? • E.g. does it matter where the gene is placed?

23. Problems With SynBio • If we continue our book analogy, writing a book is far harder than copying and pasting text from other publications (which is why plagiarism is so bad among students;) • For example: does the placement of the word in a sentence matter? • In sentence: does the matter of the example in a of matter placement? • Obviously, it does. • The same is very true for genes – we can’t just lob it in the genome somewhere – the placement of a gene will very much affect how effectively the gene is expressed.

24. More Problems • Another problem with SynBio, as well as any modified genetics, is that modified organisms can lose their competitiveness as they become more and more modified. • Many modified organisms lack the ability to compete to the extent that they cannot function outside of a lab setting. • Obviously this would be a problem for the industry. • There is no standard way to modify an organism and make it competitive and functional outside of a lab. • This makes trial and error a big part of this process.