Embedded Files
STUDENT HANDOUT:
STUDENT HANDOUT:
LABORATORY REPORT FORM (PRE-LAB PREP):
LABORATORY REPORT FORM (PRE-LAB PREP):
INTRODUCTION:
INTRODUCTION:
From the simplest bacterium to a complex multicellular organism like yourself, a fundamental characteristic of life is the presence of a genetic code in the form of DNA. DNA is a double helix molecule (Figure 1). The backbone is made of alternating deoxyribose sugar and phosphate groups, while nitrogenous bases connect the two strands together via hydrogen bonds in the middle.
In this activity, you will perform the first step necessary in most biotechnology applications - obtain the DNA! You may be aware of direct-to-consumer genetic tests; perhaps you have even voluntarily donated your DNA to be sequenced already! What happens after you provide your mouth swab in the mail? This lab will provide you with an understanding of the first step involved in this process. Extracting DNA from its protective cellular environment involves a few simple substances that rely on the hydrophilic and hydrophobic properties of the molecules within the cell.
The human genome is contained within every cell, barring a few exceptions such as mature red blood cells. While microbial DNA is not encased in a nucleus like your own eukaryotic cells, all life contains a cellular membrane with the same phospholipid bilayer. Whatever the source of the desired DNA, the first step is to break down, or lyse, the cell membrane. In this lab, the properties of the phospholipid will aid in its own destruction. The molecule is amphipathic, meaning that one part of the molecule is hydrophilic (polar) and attracted to water, while the other part is hydrophobic (non-polar) and repels water. The cell membrane stays intact because the hydrophilic ‘heads’ of the two-layered structure each face water while the hydrophobic ‘tails’ are shielded on the inside of the membrane. However, when this organization is disrupted, the bilayer can rupture. In this lab, the lysis solution contains soap which is also an amphipathic molecule. When mixed with cells, the phospholipids of the membrane reorganize and cluster with the soap molecules, spilling out the contents of the cell.
Alcohol and density will aid in the final isolation of DNA from the cellular debris. The final step requires the tube to sit undisturbed for ten minutes, as the heavier organelles and proteins settle in a tangle towards the bottom. The sugar phosphate backbone of the nucleic acid gives it polar properties. . Ethanol has both polar and nonpolar properties and thus DNA is less soluble in this solvent. Within a few minutes, the DNA will precipitate. You will see the white wispy strands of your genetic code become visible!
OBJECTIVES:
OBJECTIVES:
Understand how the structural properties of cellular components interact with various solutions.
Extract DNA from a sample of saliva.
PRECAUTIONS:
PRECAUTIONS:
Be certain to wear goggles at all times.
Do not cross-contaminate stock solutions; use fresh pipette with ethanol.
Store the final product in a cool, dark place.
If the room you are working in does not allow food/drink, complete steps 2 and 3 of the procedure out in the hallway.
MATERIALS (Work individually or in pairs):
MATERIALS (Work individually or in pairs):
Sports drink
Small cups
Test tube rack
15 milliliter graduated tubes (Falcon tubes)
2 pipettes per student
Dish soap (lysis solution)
Ice-cold ethanol (70%)
1.5 milliliter Eppendorf tube
String (optional)
PROCEDURE:
PROCEDURE:
Before beginning the experiment, read the entire procedure. Then complete the Microbiology Laboratory Report Form by answering the pre-lab questions.
Swish 2 milliliters of sports drink for 1 minute in your mouth.
Transfer the sample to a 15 milliliter Falcon tube.
Using the plastic pipet, SLOWLY measure 2 milliliters of lysis solution & add to Falcon tube with your sample.
Cap the tube and GENTLY invert 5 times.
Allow tube to sit 2 minutes undisturbed.
Holding the tube at an angle, SLOWLY add ice-cold ethanol to the tube until the volume reaches the 12-13 milliliter mark.
DO NOT MIX.
Allow tube to sit for 10 minutes undisturbed.
Obtain a 1.5 milliliter Eppendorf tube and a string.
Very SLOWLY transfer the white, wispy DNA from the Falcon tube to Eppendorf tube using a plastic pipet.
Close the tube and connect it to the string to wear around your neck (if you so wish to).
OBSERVATIONS/CONCLUSIONS:
OBSERVATIONS/CONCLUSIONS:
What observations can you make about your isolated DNA?
QUESTIONS:
QUESTIONS:
Do you think your results would have been different if you had used a different organism, such as a plant or a mushroom? Why or why not?
A student arrived late to class and simply spit into the tube to catch up with the rest of the class at step 3. Explain why their results did not yield the same amount of DNA as their classmates.
What can scientists do with DNA? Describe 2-3 scientific purposes for extracting DNA.
KEY TERMS:
KEY TERMS:
amphipathic
DNA
hydrophilic
hydrophobic
lysis
non-polar
precipitate
polar
Page updated
Report abuse