1. Synthetic Biology: Genetic Transformation by Steve Post Life Technologies / IISME Summer 2011

2. Synthetic Biology: What is Synthetic Biology? Synthetic Biology The design and construction of new biological functions and systems not found in nature. Synthetic Biology is an expansion of biotechnology, to design and build engineered biological systems that: – process information, manipulate chemicals, fabricate materials and structures, produce energy, provide food, and maintain and enhance human health and our environment.

3. Synthetic Biology: Which organisms are best to transform? One of the most commonly transformed organisms is bacteria (small, simple chromosomes, grow quickly, divide by cloning, easy to manipulate) E. Coli bacteria will grow billions of offspring in colonies overnight

4. Synthetic Biology: What do you want to create with DNA? Genetic Transformation allows scientists to modify and create genes with nearly endless possibilities. Glow in the dark E. Coli Bacteria

5. Synthetic Biology: What is genetic transformation? Transformation The insertion of new DNA into a host cell to give the cell the ability to produce new proteins and new traits. Transformation can be used for: – Agriculture: plants become more resistant to pests, frost, drought, etc – Bioremediation: bacteria can be transformed to have genes that enable them to digest oil spills – Production of medicine: bacteria can be transformed to produce proteins like insulin to help diabetics.

6. Synthetic Biology: What is genetic transformation? Transformation Requirements: Plasmid Vector DNA with Gene Recombination Insert into Bacteria Cell

7. Synthetic Biology: How do you insert genes? Vector The vector is the method used for inserting genes into a cell during transformation. – (Example: Viruses are vectors) One of the most simple and common vectors in synthetic biology is a bacterial plasmid. Plasmids are small, simple rings of DNA that can be easily manipulated.

8. Synthetic Biology: How do you use a Plasmid as a Vector? Bacteria cells have a chromosome and one or more small circular rings of DNA called plasmids. Plasmids can be replicated and pass from one bacteria to another. This occurs naturally to help bacteria adapt to new environments. Scientists can simulate this process by opening the bacteria cell membrane and insert a genetically engineered plasmid as a vector.

9. Transformation with Plasmid Vector How do you use a Plasmid as a Vector? Bacteria cells divide by cloning themselves, so all their billions of offspring have the identical transformed genes.

10. Synthetic Biology: So what traits should we give our bacteria? Why not a green glow trait? Green Fluorescent Protein glows bright green. Fluorescence is created when a substance absorbs one wavelength of light and emits a lower wavelength.

11. Synthetic Biology: What is Green Fluorescent Protein? Green Fluorescent Protein (GFP) Green fluorescent protein (GFP) is a protein composed of 238 amino acids that exhibits bright green fluorescence when exposed to UV light. GFP first isolated from a jellyfish GFP is used a marker gene : GFP is grouped with other genes in transformation, and the presence of green fluorescence from a cell is used as an indicator of successful transformation.

12. Synthetic Biology: Where did GFP come from? GFP was first isolated from the jellyfish Aequorea victoria that glows in dark environments. The gene that codes for GFP is often used in genetic engineering as a visible sign of success.

13. Synthetic Biology: What can you do with GFP? GFP is used an success indicator for desired genes. Bacteria colonies that glow green under UV light show successful transformation. Colonies that do not glow show errors or mutations. Colony #1 Colony #2 Colony #3 Colony #1 No Glow! Colony #2 Colony #3 Normal LightUV Light

14. Synthetic Biology: What can you do with GFP? Glowing colonies can be isolated and cloned to make sure every bacteria cell has the exact genes that the scientist wants. Colony #1 Colony #2 Colony #3 Colony #1 No Glow! Colony #2 Colony #3 Normal LightUV Light

15. Synthetic Biology: What can you do with GFP? Mutations of GFP create a variety of colors Bacteria art using the mutant GFP color pallet.

16. Synthetic Biology: What can you do with GFP? Bacteria are not the only GFP species… “Glo Fish” are Zebra fish engineered to be fluorescent. Glo Fish were outlawed in California in 2004. Normal Zebra Fish “Glo Fish” GFP Zebra Fish

17. Synthetic Biology: How do you transform a multi-cellular organism? To transform a multi-cellular organism, scientists manipulate the gametes or zygote. Cell division copies the genes during development, so every cell in the organism has the modified genes.

18. Synthetic Biology: What else can you do with GFP? Fluorescent Bunnies??

19. Synthetic Biology: What can you do with GFP? Fluorescent Bunnies = YES! Eduardo Kac claims he made a GFP rabbit as artwork. In fact, many rabbit embryos were transformed by scientists. The famous glowing rabbit’s name was Alba, she was an albino rabbit that glows under UV light.

20. Synthetic Biology: What can you do with GFP? Many species have been transformed with GFP, here are more realistic photos of GFP mice:

21. Synthetic Biology: Is there anything that shouldn’t glow green?