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Restriction Endonucleases

Restriction Endonucleases. David Peterson. Structure of DNA and RNA. Information Flow in Cells. DNA RNA Protein. Transcription. Translation. Amino acids combine to make peptides and proteins. Enzymes. Catalyze reactions Most are proteins.

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Restriction Endonucleases

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  1. RestrictionEndonucleases • David Peterson

  2. Structure of DNA and RNA

  3. Information Flow in Cells DNA RNA Protein Transcription Translation

  4. Amino acids combine to make peptides and proteins

  5. Enzymes Catalyze reactions Most are proteins

  6. Restriction endonucleases cut DNA at specific sites X174 DNA HinfI HaeIII

  7. Restriction Enzymes Aid in DNA Cloning

  8. Madam, I’m Adam. Gary knits a stinky rag. Harass sensuousness, Sarah. Go hang a salami, I’m a lasagna hog. Palindromes

  9. Most Restriction Enzymes Recognize Palindromes 5'-GATATC-3' 3'-CTATAG-5'

  10. EcoRV Bound to DNA www.pdb.org 1AZ0

  11. Phage l 48,512 bp dsDNA virus that grows on E. coli. Life cycle ~ 30 min at 37°C typical burst ~ 100-300 particles

  12. Bacteriophage infection: 1 → 300 → 10,000 → 3,000,000 → 109! 30’ 30’ 30’ 30’ Phage DNA Bacterium 30 min Typical phage infection cycle: 1. Phage DNA penetrates bacterial envelope 2. Bacterial DNA is disrupted & phage DNA is replicated 3. Phage structural proteins are synthesized 4. Heads, tails, fibers made and assembled into particles 5. Cell lysis, release of ~ 100 – 300 phage particles

  13. Bacteriophages Make Plaques

  14. Werner Arber Dan Nathans and Hamilton Smith Nobel Prize in Physiology or Medicine 1978

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