WET LAB DNA Barcoding: From Samples to Sequences - Northwest ...

WET LAB
Lab 3: Analyzing PCR Results
with Agarose Gel Electrophoresis
Teacher Preparation
• Load the classroom computer with the Wet Lab PowerPoint slides.
• Make copies of the Student Handout—Analyzing PCR Results with Agarose
Gel Electrophoresis, one per student. These handouts are designed to be
reused as a class set; students write answers to questions and take notes on a
separate sheet of paper or in their lab notebook.
• Teachers may wish to have their students write out the lab procedures before
the activity in their lab notebooks or on a separate piece of paper as a “prelab”
exercise, which can be used as an “entry ticket” to class. During the
lab, students may check off steps as they complete them and/or describe and
draw pictures of their observations.
• If students are not familiar with agarose gel electrophoresis, queue your
computer to one or more of the following tutorials:
‣ “How to Make and Run an Agarose Gel (DNA Electrophoresis)” by
labtricks.com (4:54 min) includes how to calculate percent agarose, make
and pour an agarose gel, a close-up of sample loading on the gel, and
running the gel: http://www.youtube.com/watch?v=2UQIoYhOowM.
‣ “Loading a Gel for Electrophoresis” by Carolina Biologicals (5:46 min)
includes different types of pipettes used and a close-up of the proper
techniques to load a pre-poured agarose gel. [Note: The first 2:50 min of
the video covers loading a gel with a micropipette.]: http://www.youtube.
com/watch?v=h06rz8rcZpw&feature=related.
‣ “DNA Electrophoresis Sample Loading” from Greg Peterson, Coordinator of
Biotechnology, Kirkwood Community College, Math/Science Department,
includes pouring the agarose gel, preparing the DNA samples for loading
on the gel, and close-ups of the “do’s and don’ts” of loading DNA
samples on gels, including a number of common mistakes. [Note: The
first 1:00 min of the video moves quickly, and the voice-over does not
precisely match the video content; however, the sample loading examples,
beginning at 1:04 min, are extremely helpful. If this video is not shown
to students, teachers may want to review it to anticipate and evaluate
common gel loading problems encountered by students.]: http://www.
youtube.com/watch?v=tTj8p05jAFM&feature=related.
• Set up student work stations by distributing supplies and reagents needed
for each group, as described above under Materials. It is suggested that
reagents be aliquoted for student groups and labeled as described in
Student Handout—Analyzing PCR Results with Agarose Gel Electrophoresis.
This information is found in Teacher Resource—Aliquoting DNA Barcoding
Reagents for Labs 1–4.
• Visualizing DNA Gels with Fast Blast DNA Stain. DNA gels can be
stained quickly (i.e., in less than 20 minutes) with 100X Fast Blast DNA
stain or overnight with 1X Fast Blast DNA stain. Students are instructed in
Student Handout—Analyzing PCR Results with Agarose Gel Electrophoresis to
[Note: Students often confuse the tubes
for the loading dye and the molecular
weight standard. Both are in small, 1.7
ml microfuge tubes, and both are blue.
Teachers may wish to only set out the
loading dye, and hand out the molecular
weight standard after all samples have been
prepared. If students have no bands in their
molecular weight standard lane, they may
have loaded dye instead of the molecular
weight standard. If there are many bands
in their sample wells, they may have mixed
their samples with molecular weight
standard instead of loading dye.]
[Note: Fast Blast DNA Stain can be
re-used up to 7 times.]
Wet Lab – DNA Barcoding: From Samples to Sequences
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©Northwest Association for Biomedical Research—Updated October 2012