Transfection of DNA into Eukaryotes

1. Transfection of DNA into Eukaryotes “Mom, please, may I have a glowing fish, too?”

2. Although glowing fish are fun, the procedure used to make them is very useful to scientists!

3. We are going to make living cells glow

4. Transfection • Delivery of foreign molecules (DNA) into eukaryotic cells • Cell begins to read the piece of DNA and produce the protein encoded on it

5. How can we get DNA into a cell? • Inject it • Poke holes in the membrane without destroying the cells • Create a complex of DNA plus some substance that will go into the cell

6. We will use Cationic liposomes • It creates an artificial membrane • Membrane formed around pieces of DNA • Cell membrane will fuse with this artificial membrane and bring the DNA into the cell Liposomes are also be useful for drug and cosmetic delivery

7. Liposome-DNA complex taken up by the cell by fusing with the cell membrane

8. Can change an entire animal or just a culture of cells • View with fluorescent microscope • Use blue or ultraviolet light

9. GFP • Green fluorescent protein • Gives luminescent jelly fish their greenish glow

10. Plasmid Vector • GFP is a marker protein on a plasmid vector • A plasmid is a piece of DNA coding for a specific protein • If the cell read the DNA and produces the protein, it will also produce GFP. • That particular protein will glow • If there is enough of that protein, the entire cell will glow

11. What can we do with glowing cells? • Determine distribution of protein • Track movement of protein • Track movement of cells • VERY useful for scientists!