Ap Biology Chapter 20 Notes

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Chapter 20: DNA Technology
Biotechnology: Use of living organisms to perform tasks.
* Wine & cheese
* Selective breeding
* Antibiotic production
* Recombinant DNA
Restriction Enzymes
* Bacterial enzymes: cut up foreign DNA
* Specific: only but at recognition sequences
* Palindromic: cut at the same base sequence on each strand, but in the opposite direction * The exposed bases provide “sticky ends”
* H-bond to compliment bases of segments cut with same restriction enzyme Recombinant DNA
1. Isolate DNA from 2 sources
* Bacterial plasmid w/2 genes
* ampR(resistance to ampicillin)
* lacZ(makes β galactosidase)
* Human cell
2. Treat both w/ the same restriction enzyme
* Recognition sequence cuts through lacZ gene in plasmid
3. Mix all DNA fragments with plasmids.
* H-bonds bt/n bases temporarily hold “sticky ends” together 4. Add DNA ligase: creates permanent covalent bonds
5. Reintroduce plasmid to bacteria via transformation
6. I.D. recombinant cells by
* Ability to grow in
* Inability to digest x-gal(the colonies will be white, not blue) X-gal -------------------------------- (β galactosidase)---------------------> Blue product (molecular mimic of (white)lactose) lacZ gene

7. Each bacterium replicates to form thousands of clones
* Many different human DNA fragments have been cloned
* -------------------------------------------------
How do you find the one “of interest”?
Nucleic Acid Hybridization
* Synthesize a Nucleic Acid Probe
* Single strand DNA or RNA (radioactive)
* Complementary to the gene of interest
* Synthesize on the base of:
* Base sequence of the gene (at least part of it)
* AA sequence of the protein
* Denature the DNA with heat or chemicals
* The probe will H-bond to the appropriate gene
Genomic Library
* Complete set of recombinant clones (includes all fragments) * Stored in a collection of phage clones (virus)
Sources...
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