Simple Thymus DNA EXtraction
Mary Jo Osborn
Sacred Heart Griffin High School
This procedure combines ideas taken from several sources including The Onion Lab: DNA Spooling by Judy Brown, Down and Dirty DNA Extraction by Rod Bost, and an unattributed hand-out called DNA Extraction received from Dr. George Kieffer, University of Illinois.
- Calf Thymus (also called sweetbreads) may be cut into pieces and frozen for future use.
- Isotonic sucrose solution ( 106.1 grams table sugar in 1 liter of water )
- Cheese cloth
- Detergent solution ( 100 mL dishwashing liquid [ I've used Dawn], 15 grams NaCl [table salt works fine]: add distilled water to make a final volume of 1000 mL)
ethanol (95%) dispensed into 3-5 mL amounts into screw top test tubes or small containers and stored in a freezer.
One container per student group.
2 test tubes per student group
- inoculating loops or thin wires bent to form loop at the end for drawing up DNA from test tube
As much as possible, you would like to work with cold materials and solutions. When the alcohol is removed from the freezer it should be carried on ice into the classroom.
- To break open the thymus cells: Add a small piece of thymus (slightly smaller than an egg) to the blender with enough sucrose solution to make a thin milkshake consistancy. Blend until thymus is broken up. Kids think this is cool and gross to watch!
- Pour the blended thymus through a funnel which is lined with four thicknesses of cheese cloth. The filtrate collected below the funnel will contain many nuclei which can be observed under the microscope. This step removes gristle and other stuff.
- Pour about 2 mL of the nuclei suspension collected from step 2 into a clean test tube for each student group.
- Pour about 2 mL of detergent solution into another clean test tube for each group.
- Students pour detergent solution into test tube of nuclei suspension and gently shake tube from side to side to mix. Do not shake enough to get foamy! In this step the detergent will break open the lipid membrane of the nuclei and release DNA. The salt in the solution will help neutralize the negative charges on the DNA molecules so they will precipitate.
- Slowly pour COLD ethanol down the inside of the test tube containing the nuclei/detergent mixture so that a layer of ethanol forms on top of the mixture. DNA will come out of solution at the place where the two layers meet.
- Use clean wires to dip into layer where the two solutions meet and stir gently. Students will be able to lift out masses of DNA. I have had students pull gently until they lifted about 2 meters of long chain molecules out.