DNA Extraction: A Laboratory Using Common Ingredients

DNA may be extracted from cells using common household chemicals. The procedure noted below will extract DNA from yeast cells. The low cost and availability of the reagents as well as the simplicity of this laboratory procedure make it a useful demonstration of the practice of biotechnology. The crude nature of this procedure and the general specificity of the reagents prevent the extracted DNA from "spinning out" of the final phase. The time required for this laboratory exercise is approximately one class period. The procedure has been presented in the form of a recipe, common kitchen instructions

ARTICLE:

DNA may be extracted from cells using common household chemicals. The procedure noted below will extract DNA from yeast cells. The low cost and availability of the reagents as well as the simplicity of this laboratory procedure make it a useful demonstration of biotechnology. The crude nature of this procedure and the general specificity of the reagents prevent the extracted DNA from "spinning out" of the final phase. The time required for this laboratory exercise is approximately one hour and fifteen minutes. The procedure has been presented in the form of a recipe, common kitchen instructions.

Ingredients: (Materials and Equipment):
beaker, 250 ml
water, 100 ml (tap water is acceptable)
yeast, 1/2 package of Fleischmann's All Natural Yeast
or
2 oz. of baker's live yeast (brick form)
Woolite or Dawn detergent, 5 ml
Adolph's 100% Natural Tenderizer-Unseasoned (contains the enzyme, papain), 3gm.
Alcohol, 100 ml. (95% Ethanol or 50% Isopropyl or 95% Methanol)
Baking Soda, 500 mg. (Sodium Bicarbonate 500 mg/100 ml water produces a pH 8 buffer solution).
thermometer (0 - 100 C)
glass stirring rod and/or pipette
watch glass or Petri dish

OVERVIEW: The yeast will be suspended in the water and subjected to the action of the enzyme from the tenderizer. This "cooking" will break down the cellular structure of the yeast cells releasing the contents of the cells into the solution. This process will be aided by heat, the addition of some surfactant (detergent) and the buffering of the baking soda to hold the pH near 8. Once "cooked" the free DNA may be separated from the "soup" using the alcohol to selectively precipitate any DNA fragments. This procedure tolerates many inconsistencies, however the yields of DNA may vary with any change in methods. Yeast DNA extracted with this method, usually, will not "spool" out of solution in long filamentous strands, as occurs with other extraction methods. This may be due to the presence of some DNase either in the meat tenderizer or introduced through contamination. Any DNase activity tends to break the long DNA strands into smaller pieces.

Step 1: Prepare the yeast by dissolving the brick of live yeast in 100 ml of warm water (50-60 C.) If you are using the equivalent "dry" yeast, let the solution stand until all of the yeast granulars are soft. (While they remain dry, you will feel a "grittiness" between your fingers if you feel a drop of the solution. Once they have dissolved, a drop of this solution will feel smooth between your fingers.)

Step 2: Add 5 ml of surfactant (detergent) to the yeast and maintain a temperature of 50 C. for 5 minutes. DO NOT OVERHEAT. DNA will denature completely at about 80 C. Keep the temperature at 50 C. The detergent will remove some of the lipids (fats) that make up the cell walls of the yeast cells. This step will "open up" the yeast cells so that the enzyme may get to the proteins within the cells.

Step 3: Add the 3 gm of tenderizer and the 500 mg of baking soda. Stir until these dissolve. Then continue cooking at 50 - 60 C. for 20 minutes. The enzyme papain (a proteolytic enzyme) within the tenderizer will break down many of the proteins that bind the DNA within the yeast cells. The Sodium bicarbonate acts as a pH buffer and holds the pH close to a value of 8. This pH allows the enzyme to degrade many of the proteins yet leave the DNA itself somewhat untouched.

Step 4: Allow the entire solution to cool to room temperature (20 C.) This entire solution may be stored in a refrigerator overnight for use the following day.

Step 5: Separate some of the solution into smaller test tubes with about 1 ml of solution per each test tube. You only need one test tube per student to demonstrate this process.

Step 6: Gently layer 1 ml of cold alcohol on top of the lysed solution. This may require some practice. Pour the alcohol slowly down the inside of a tilted test tube. It will flow on top of the layer of lysed solution. DO NOT SHAKE.

Step 7: Hold the tube upright and observe the interface of the alcohol and the solution. Slowly insert the glass rod into the liquids and gently stir with a constant motion. The interface between the aqueous solution of lysed yeast and the alcohol should enlarge. Do not mix so hard that these two liquid phases emulsify. As the free DNA comes into contact with the alcohol a precipitate of DNA will form at the interface of the fluids. If the DNA is intact and in the form of long strings, these will adhere to the glass rod. (This procedure tends to break down the long linear strands of DNA.) If the enzyme and heating have broken the DNA into smaller fragments, these will appear to "float" within the alcohol layer. The alcohol containing DNA may be removed with a pipette and the alcohol allowed to evaporate by placing the DNA/alcohol suspension in a watch glass or Petri dish.