Gene Therapy Yerr baby!!!!

The main advances in medicine Improved sanitation systems Surgery with anesthesia Vaccines and antibiotics And the fourth will be Gene Therapy

What is gene therapy? The selective delivery of genes into a patients cells create missing proteins replace defective disease causing genes Over 4,000 conditions are caused by damage to a single gene, many others by several genes

Gene therapy techinques Ex-vivo “outside the body” adding blood cells with new DNA to the blood stream but has a limited time span Using bacteria like E.coli to produce billions of copies of a human gene Using viruses to carry the gene to specific cells in the body

What do you need? A cutting enzyme called a Restriction enzyme It cuts foreign DNA that enter bacteria, e.g. Eco. R1 A circular DNA molecule from a bacteria called a Plasmid The E. coli bacteria has dozens of different plasmids A section of the human DNA containing the gene you need An enzyme to join the bacterial DNA and human DNA called ligase

Restriction Enzymes Found in many bacteria A defense mechanism which cuts foreign DNA The DNA is not cut randomly, but at specific sequences called Recognition sequences The restriction enzyme Eco.R1 found in strain C, E. coli bacteria They don’t make straight cuts, but produce sticky ends These sticky ends can rejoin by forming hydrogen bonds and the sugar-phosphates rejoining with the help of the enzyme ligase The DNA produced by restriction enzymes cutting is called gDNA We know of 200 different restriction enzymes

Plasmids The Plasmid pSC101 was isolated from E. coli. It is useful for gene therapy because it has only one sequence of GATTC in its entire molecule Plasmids can be used to replicate DNA segments of up to 4,000 base pairs in length For longer length viruses must be used.

Using Bacterial Plasmid Cut the Bacterial Plasmid using restriction enzyme called Eco R1

This produces a hole in the circular Plasmid DNA Eco. R1 only cuts at the sequence GAATTC This produces two identical ends

Plasmid Also treat human DNA with Eco.R1 and you get a section of DNA containing the gene you want with two complementary ends as well! Short sequence of DNA with sticky ends approaches a Plasmid. Human DNA

Bacterial DNA Human DNA The newly completed piece of DNA is called Recombinant DNA

Using this Recombinant DNA Gene cloning can now be used to produce millions of copies

A Plasmid with human DNA is put back into a E. coli The bacteria expresses the DNA. The bacteria then divides to produce millions of copies bacteria and human gene.

A Plasmid cut by a restriction enzyme Foreign DNA spliced into Plasmid by Plasmid enters bacterial cell Segment of DNA approaches DNA ligase joins phosphates and sugars

Using viruses Viruses can be used to transfer large pieces of human DNA to a location in the Human body. Viruses are made up of a molecule of nucleic acid and a protein coat. Their nucleic acid codes for protein coats, replication, enzymes to break-in and out of particular cells. You remove the part of the nucleic acid that does not code for the above functions and add foreign DNA. The virus can then be put into the blood stream and it will enter the cell it is designed to attack and transfer its code into that cell.

Some current uses of gene therapy and later cosmetic uses Some current uses & trials of gene therapy Introduction of genes for new blood vessels for clogged arteries Introduction of genes into the blood stream that interferes with the replication of HIV Human growth hormone for Dwarfism Insulin production Glowing mice Replacing the cytoplasm in eggs to produce healthier eggs Some current uses of gene therapy and later cosmetic uses

The future? Injecting genes into the blood stream which are then carried to the target cells by viral carriers where they unload their genetic material. This is then used by the body to produce proteins that fight the disease. For the next decade gene therapy will only be used on somatic cells (only effect the patient and not their offspring) You and bioethicists will have to decide the future!