1. Polymerase Chain Reaction PCR
Use to exponentially amplify discrete regions of DNA.
Need to know the DNA sequence of two short regions, not too far apart, but don’t need to know the sequence of the entire region to be amplified.
Make two oligonucleotide primers (ea. ~20 bases long), complementary to opposite strands at the ends of the region to be amplified.

3. - Gel Electrophoresis +
Separates molecules (DNA, RNA, or proteins) on the basis of size.
Gels: Agarose or Acrylamide
DNA gels:
- Load DNA samples into ‘wells’ at one end of gel.
- Apply electric current through gel.
- DNA pushed toward + electrode.
- Smaller molecules travel faster through gel, resulting in separation of DNA fragments on basis of size - +
Ethidium Bromide binds to DNA & fluoresceses when exposed to UV

4. Restriction Mapping Digest DNA with restriction enzymes:
17 kb
Restriction Mapping
Digest DNA with restriction enzymes:
Single enzyme digests
& Double enzyme digests
(two enzymes together)
Determine sizes of restriction
fragments
Deduce locations of restriction enzyme
recognition sites

5. Soak gel in NaOH
Run DNA on gel
Southern Blots
Determine whether two DNA fragments contain same or similar DNA sequences.
DNA transferred to filter paper
= Southern blot
Hybridize Southern blot with radioactive probe (same as in library screening)
Wash filter & expose X-ray film
Only restriction fragments on Southern blot that contain sequences complementary to probe will show up as bands on autoradiogram

6. Transfer DNA from gel to piece of DNA binding filter paper

7. Mapping Chromosomal Rearrangement Breakpoints
Genomic Southern blots
Reciprocal Translocation
Fig. 9-13