Transcript pGLO

Gel Electrophoresis Lab
Analysis of Precut Lambda DNA
Restriction Enzymes
Our lambda virus DNA
samples have been precut
with three different
restriction enzymes.
L – lambda DNA
P – PstI lambda digest
E – EcoRI lambda digest
H – HindIII lambda digest
Running the Gel
• The phosphate groups in
the sugar-phosphate
backbone of DNA are
negatively-charged.
• Molecules move through
the gel at different rates,
determined largely by
their size and charge.
Shorter molecules move
through the gel faster
than longer molecules.
Reasons for Each Step
 Loading Dye
Two sizes of blue molecules, small
and large, help visualize movement
of sample across gel.
 Heat DNA Samples
Denatures double stranded DNA to
produce single strands that more easily
move through gel.
Reasons for Each Step
 Tap Tubes on Table Top
moves sample to bottom of tube for collection
 Run the Gel in Chamber
electrical field pulls negatively charged DNA through
the gel, with distance moved proportional to size
 Stain Overnight
stain attaches to DNA within gel, making bands of
DNA visible
Lab Tips
• Follow the “How to Use a Micropipet” instructions at
your station.
– When finished, eject the tip into the disposal at your station.
• Carefully handle the DNA samples!
• Be sure your gel is well-side up and fully
submerged in the buffer solution.
• Hold the micropipet just over the
well. Be careful not to puncture
the bottom of the well with the
micropipet!
• Use tape to label which gel is yours (top or bottom).