Gene therapy 1


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Gene therapy 1

  1. 1. Gene Therapy Introduction Pages 1-11
  2. 2. Gene Therapy <ul><li>Purpose: </li></ul><ul><ul><li>Introduce a functional gene into a person who lacks this gene </li></ul></ul><ul><ul><li>Gene therapy is potentially useful for treating recessive diseases </li></ul></ul><ul><ul><li>Parents are heterozygotes </li></ul></ul><ul><ul><li>Patients lack a particular enzyme </li></ul></ul><ul><ul><li>Gene therapy is used to replace the missing enzyme </li></ul></ul>
  3. 3. Gene therapy <ul><li>The first disease that gene therapy was used for was Severe Combined Immune Deficiency (SCID) </li></ul><ul><li>In this disease, there is a lack of the enzyme adenosine deaminase (ADA) </li></ul><ul><li>Gene therapy is used to give a patient a functional ADA gene </li></ul><ul><li>Introduce the gene into bone marrow cells and the patient can then produce WBCs throughout their life and produce the enzyme </li></ul>
  4. 5. What diseases should gene therapy be used for? <ul><li>Diseases must result from the defect in a single gene </li></ul><ul><li>Gene must have been identified and purified </li></ul><ul><li>Cells affected by the gene defect must be accessible for gene therapy </li></ul><ul><ul><li>Problems in treating neurological disease </li></ul></ul><ul><ul><li>Must get past the blood brain barrier </li></ul></ul><ul><li>Diseases chosen usually have few effective treatments </li></ul>
  5. 6. What cells should be targeted to be altered? <ul><li>Gene therapy is for altering somatic or body cells and not germ cells </li></ul><ul><li>This treatment will be used to correct a defect in an individual but the correction is NOT passed on to subsequent generations </li></ul><ul><li>Gene therapy is used to treat diseases and not alter characteristics of the next generation </li></ul>
  6. 7. Transcription and Translation <ul><li>DNA codes for the gene that you want to insert </li></ul><ul><ul><li>Insert the gene for ADA to treat SCID </li></ul></ul><ul><li>If gene therapy is successful, then the ADA enzyme will be made </li></ul>
  7. 9. <ul><li>SCID - ADA </li></ul><ul><li>Lung cancer - p53 tumor suppressor gene </li></ul><ul><li>Cystic fibrosis - Cystis fibrosistransmembrane conductance regulator (CFTR) </li></ul><ul><li>Leukemia - Interleukin 2 (IL-2) </li></ul><ul><li>Look at table 2, pages 5-7 </li></ul>Different diseases can be treated by introducing different genes
  8. 10. How do you make the recombinant DNA? <ul><li>Vector </li></ul><ul><ul><li>Plasmid </li></ul></ul><ul><ul><li>Place to incorporate the foreign DNA </li></ul></ul><ul><ul><ul><li>The DNA that codes for the missing enzyme </li></ul></ul></ul><ul><ul><li>A marker gene - antibiotic resistance gene </li></ul></ul><ul><ul><ul><li>Determine if the vector entered the cell </li></ul></ul></ul><ul><li>Foreign DNA </li></ul>
  9. 11. Steps in making recombinant DNA <ul><li>Need DNA from 2 different sources </li></ul><ul><ul><li>In this class, we will transfer DNA from one plasmid and put it into another plasmid </li></ul></ul><ul><li>Cut DNA with restriction enzymes </li></ul><ul><ul><li>Create staggered or blunt end cuts </li></ul></ul><ul><li>Match the DNA from the 2 different sources and let them combine </li></ul><ul><li>DNA ligase seals the 2 pieces together </li></ul>
  10. 13. DNA structure
  11. 15. When performing cloning, how do you find the gene you are interested in? <ul><li>Get a gene library </li></ul><ul><ul><li>Collection of different genes </li></ul></ul><ul><li>Use a PROBE to find your gene in this gene library </li></ul><ul><li>A probe is complementary to your gene of interest </li></ul>
  12. 16. How is the PROBE used? <ul><li>Gene sequence = AGCTGGAC </li></ul><ul><li>Probe = TCGACCTG </li></ul><ul><li>Heat is used to separate the two strands </li></ul><ul><li>Probe is added to the gene sequence </li></ul><ul><li>Probe is labeled at one end with a radioisotope so it can be detected if it binds to the gene </li></ul>
  13. 18. How is the PROBE used? <ul><li>Locate a functional ADA probe to perform gene therapy for SCID </li></ul><ul><li>Use a gene library from someone who DOES NOT have SCID </li></ul><ul><li>Use a probe specific for the ADA sequence </li></ul><ul><li>Isolate the DNA to be used in gene therapy </li></ul><ul><li>Probes are also used in Southern Blots, Northern Blots and Western Blots </li></ul>