PHRD5025 Lecture 16 - Recombinant DNA Technology

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PHRD5025 Lecture 16 - Recombinant DNA Technology
2013-11-18 02:58:57
Recombinant DNA Technology

Recombinant DNA Technology
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  1. blotting for mRNA analysis
    Northern blotting
  2. spontaneous association of complementary nucleic acid chains
  3. 3 steps of PCR
    • 1) denaturation
    • 2) annealing
    • 3) elongation
  4. type of PCR used to detect mRNA
    reverse transcriptase PCR
  5. purpose of qPCR (real-time PCR)
    quantify mRNA
  6. type of blotting used for DNA analysis/fingerprinting
    • Southern blotting
    • (involves PCR)
  7. assay that allows for the simultaneous quantification of all mRNAs in a sample
    DNA microarray (Chip)
  8. 5 steps in microarray assay
    • 1) nucleic acid isolation
    • 2) denaturation
    • 3) probe annealing
    • 4) detection
    • 5) quantification
  9. evaluates all the proteins expressed in a cell
  10. differences found in a population
  11. genetic differences between individuals at a location that is represented in <1% of the population
    allelic variations
  12. allelic variation at a position that occurs in a human population at a frequency >1%
    genetic polymorphism
  13. 2 steps of drug metabolism
    • 1) Phase I: formation of reactive intermediates by CYP450
    • 2) Phase II: conjugating enzyme systems to make intermediates more water soluble for elimination
  14. polymorphisms in the 2 phases of drug metabolism that put individuals at higher risk for cancer
    • 1) increased Phase I activity
    • 2) decreased Phase II activity
  15. treatment that aims to cure disease by correcting or replacing known genetic errors or deficiencies
    gene therapy
  16. what vector is mostly used to deliver gene of interest in gene therapy?
    viral vector
  17. first NIH sponsored gene therapy approach was aimed at this disease (bubble boy disease)
    adenosine deaminase (ADA) deficiency
  18. replication incompetent
  19. 3 major barriers for gene therapy
    • 1) identification of the genetic basis of the disease
    • 2) safe and efficient delivery of DNA to affected cells
    • 3) greater understanding of mechanisms of gene regulation