More PCR stuff flashcards exam III.txt

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Author:
rincrocci
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120656
Filename:
More PCR stuff flashcards exam III.txt
Updated:
2011-12-04 12:04:51
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pcr
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pcr
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  1. Uses for PCR based technology
    • Gene therapy
    • genotyping
    • Transgenics (GMO's)
    • sequencing
    • mapping
    • cloning
  2. Recombinant DNA technology
    • involves isolation & purification of genes
    • uses genetic molecular markers (ie gfp or zeocin)
    • makes modifications or reintroduces genes (ie lmp2a)
  3. 3 things needed for Recombinant DNA
    • 1. Restriction endonucleases
    • 2. DNA ligase
    • 3. cloning vectors
  4. Restriction cuts features
    • Recognition sites are PALINDROMES (seq reads same as reverse)
    • Sites are 4, 6, 8 bp in length
    • Cleavages generate the following: blunt ends, 5' or 3' overhangs
  5. Steps in making & selecting for recombinant DNA
    • 1. Cut foreign DNA - at Restriction enzyme site
    • 2. Transform bacteria
    • 3. Plate bacteria and see growth - if cut in between a tetR for example, will not be resistant to tet
  6. Blue-White selection
    • Utilizes lac z - which makes B-Galactosidase
    • X-Gal reacts with B-galactosidase & makes blue precipitate
    • When you plate transformed back on X-gal, the cut thru lac z will be white
  7. Restriction Maps
    • Restriction enzymes used to ORDER the recognition sites on piece of DNA
    • (do examples as in HW III #1)

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