MacGyvering Gel Electrophoresis: a Research-based Lab

Lacey HowardShearer Lab Research Group

Sumrall High School Biology I StudentsSorrell: 1st block 30, 3rd block 20Hummell: 3rd block 21

5 E’s of the Lesson Plan

1. Engage- ask students to discuss their knowledge of gel electrophoresis.

2. Explore- have the students practice using pipettors with a scale and water.

3. Elaborate- have the students perform their own gel electrophoresis experiment.

4. Explain- discuss the principles of gel electrophoresis and how it is used in my research.

5. Evaluate- have the students fill out lab worksheets with data results and summary questions.

State and National Subject Content

• Science as Inquiry– Using the scientific method

• Science and Technology– Distinguish and explain the applications of various tools and

techniques used in DNA manipulation– Steps in genetic engineering experiments – Use of restriction enzymes

• Science in Personal and Social Perspectives – Explain uses of technology in paternity testing and crime

scene evaluations.• Unifying Concepts and Processes

Materials Required

Each gel electrophoresis kit needs:• Five 9 V batteries connected in series• Two alligator clips with one red wire and one

black wire• Small Tupperware with two holes on either

end• Two wires bent through holes in Tupperware

and connected to alligator clips

Additional Materials

• 1X TAE buffer (Tris base, Acetic acid, EDTA) or 1x TBE buffer (Tris base, Borate, EDTA)

• Agarose• Loading Dye• Ethidium Bromide• Safety glasses• UV light• Surgical gloves• DNA• Erlenmeyer flask• Microwave

What is gel electrophoresis?

• Gel electrophoresis is a technique that separates pieces of DNA (or other biological molecules) by size so researchers can analyze them.

How does gel electrophoresis work?

Wells for DNA samples

- Electrode

+ Electrode

Top View

Side View

- Electrode + Electrode

Agarose gel with Ethidium Bromide Staining

Larger DNA Fragments

Smaller DNA Fragments

DNA Ladder

Molecular Weight Marker

• A mixture of different pieces of DNA of known size used to determine the size of fragments in the sample.

Staining

• DNA in the gel is invisible.• To visualize DNA, gels are stained with ethidium bromide.• A UV light is then used to visualize the EtBr.

CAUTIONS:• Ethidium Bromide is a carcinogen.

– wear gloves– correct disposal– UV light damages eyes– wear goggles– use photoimaging system

Ethidium bromide

• Ethidium bromide has a ring structure similar to the ring structure in DNA bases; therefore it intercolates between the bases.

• Shining UV light on ethidium bromide will cause it to fluoresce at visible wavelengths.

Loading Dye

• Purposes– give color to sample– give weight to sample so it will sink into well– show progress of gel migration

• Components– dyes (xylene cyanol, bromphenol blue, Orange G) weight (glycerol or ficoll)

Uses of DNA Gel Electrophoresis

• Is DNA present in sample?• Is a PCR product present?• Is the PCR product the correct size?• Where in the DNA do specific enzymes cut?• Also, a specific DNA fragment can be

recovered from the gel.

Gel Electrophoresis Start to Finnish

Biology I Students Performing Electrophoresis

Changes for the Future

• This lab requires at least an hour of preparation time.• To minimize the time students are waiting for the

experiment to run without having work to do, give the lecture while the experiment is running.

• Explain more about PCR and restriction enzymes during the lecture.

• Engage students by talking about processes they see on crime scene shows like paternity testing and DNA evidence.

• Go over proper pipetting technique and aseptic technique before lab experiment.

References

• Desktop electrophoresis lab- moving molecules http://www.accessexcellence.org/AE/WWC.1993/moving.php

• DNA gel electrophoresis http://www.colorado.edu/Outreach/BSI/pdfs/Electrophoresis%20Notes.pdf

Q & A