AP Biology
Unit 3
RFLP
Stands for Restriction How many fragments will result
when each of these alleles are
Fragment Length
digested with DdeI?
Polymorphism
3 fragments
Takes advantage of
differences in DNA
between individuals that
result in different
fragments when digested
with restriction enzymes
2 fragments
RFLP
To see RFLP, DNA is
digested with the
appropriate restriction
enzymes and run on an
agarose gel.
A Southern Blot is
performed to complete
the analysis.
Southern Blotting
A method to visualize
specific segments of DNA
usually a particular gene.
Uses radioactive probes
that bind to the specific
DNA segments
Ex. When testing for the
hemoglobin alleles, the
probe would bind to these
regions
Southern Blotting
Steps:
Soak gel in basic solution to separate DNA
strands
Transfer DNA on to a nylon membrane
(spacing of DNA is maintained)
Incubate with radioactive probe for specific
segment
Wash away unbound probe
Detect probes using x-ray film
autoradiograph
RFLP Animations
Animation #1
Animation #2
Overview of PCR
Uses different temperatures to amplify
DNA
Step 1: Separate existing DNA strands
95C (Denaturation)
Step 2: Lower temperature to allow primers
to bind to target DNA 55C (Annealing)
Step 3: Raise temperature to allow Taq
Polymerase to build DNA strand 72C
(Extension)
PCR animation
animation