Outbreak Lab: In this lab, biotech procedures will be used to
see if a sample of viral DNA is the deadly Alabama virus. The
specific technique that you will perform is a gel electrophoresis
of digested viral DNA At the end of the lab investigation, you will
have created a DNA profile. A DNA profile is an analysis of DNA
fragments to determine if it comes from a particular individual or
organism or virus. You are creating a DNA fingerprint.
Slide 2
How can you create a DNA Profile? 1.DNA Extraction remove DNA
from virus 2.Polymerase Chain Reaction (PCR) make copies of the
viral DNA in questions 3.DNA Restriction Digest use restriction
enzymes to cut DNA at given sites 4.Gel Electrophoresis separate
the fragments of viral DNA to identify it
Slide 3
Here are the three samples of digested viral DNA. The red
capped vial has the Alabama strain, the yellow capped vial has the
Missouri strain and the green capped vial contains the Pennsylvania
strain. Note the blue liquid in the red capped vial. There is 20 ul
of DNA and loading dye.
Slide 4
Prepare the gel for the electrophoresis The bottle with the
blue gel will be melted in a microwave. The blue liquid gel will be
poured into the gel rig in front of the bottle. Notice a plastic
white comb in the gel rig. This comb will form all the gel to form
wells for the samples of DNA in the previous slide.
Slide 5
Why is the gel blue? The gel is blue because a stain called
methylene blue was added to the gel when it was made. The small
bottle to the left of the large gel bottle contains the stain. This
stain will be used to stain the DNA fragments during and after the
electrophoresis. Be careful: it will stain hands and clothes.
Slide 6
This is the work area for the gel electrophoresis. Look at the
gel box that contains the blue liquid. The blue liquid is TAE
buffer. It is blue because methylene blue stain was added. The box
is hooked up to the electrophoresis electric supply which will
supply the current to move the DNA fragments down the gel. A
micropipette set to 20 ul and yellow tips are ready for
loading.
Slide 7
Gel in the gel box filled with TAE buffer Note the comb is
still in the gel. This must be removed before loading with
DNA.
Slide 8
Loading the DNA Sample The gel is being loaded with the
Missouri strain virus DNA. Other wells will be used for the Alabama
and Pennsylvania strains.
Slide 9
Below is a gel that has run. Note the blue line at the bottom
of the gel. This is the loading dye that was mixed in with the
viral DNA. It runs faster than the fragments. The fragments are
hard to see. To visualize the fragments, the gel will be
stained.
Slide 10
Visualizing the Restriction Fragments Staining the
gelsDe-staining the gels
Slide 11
The DNA Fingerprint Examine the three lanes. Do any of the
lanes exhibit the same pattern of DNA fragments? Lane Yellow and
Green have the same restriction pattern. This means that the DNA
from these two states are the same which means that the virus
infecting these two states are the same strain. Red Lane was
Alabama strain. Yellow Lane was the Missouri strain. Green Lane was
the Pennsylvania strain. Red YellowGreen