LAB 2

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Grand Canyon University *

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181

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Biology

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Dec 6, 2023

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docx

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X19A - Module 2 Lab#2 : 35 points Part A. Observing the Effects of Osmosis on Cells Part B. Wet Lab: Beet Membranes Part C. Wet Lab: Osmosis and Diffusion Part A. Observing the Effects of Osmosis on Cells Instructions: 1. Study depictions of plant & animal cells exposed to different solutions 2. Fill in data table with your observations & answer the questions that follow 3. Complete Part A of the Lab#2 worksheet. Move on to Parts B and C. Submit all parts of Lab#2 together. Part B. Wet Lab: The Effect of pH and Detergent on Biological Membranes Introduction Biological membranes are composed of a phospholipid bilayer with a mosaic of proteins embedded within it. Since protein structure is sensitive to environmental conditions such as temperature, pH, and various chemicals, these same conditions can also affect the integrity of cell membranes. Additionally, exposure to detergents can also disrupt the phospholipid bilayer. Detergents are amphipathic molecules that have both hydrophilic and hydrophobic portions, which is why they make good agents for cleaning grease. In this experiment, you will test the effect of pH and exposure to a detergent on the integrity of beet cell membranes. Plant cells contain a large central membrane-bound vacuole called a tonoplast . In red beet cells, the tonoplast contains a bright red pigment called betanin. If the beet cell tonoplast becomes damaged, betanin will leak out into the surrounding medium. You will measure the relative amount of tonoplast membrane damage caused by pH by observing the amount of betanin that has leaked out from the beet cells. In addition to betanin release, pay careful attention to the way the beet itself looks. Note that at very high pH levels, the betanin itself may become "bleached" out. Materials 1. one red beet root 2. eight test tubes 3. test tube rack 4. vinegar (strong acid) 5. baking soda solution (strong base): dissolve ~1 tsp (5 ml) of soda in ½ cup (125 ml) of water 6. liquid dish detergent 7. water 8. pH test strips 9. medicine droppers
X19A - Module 2 10. glass stirring rod (or long thin straight stirring utensil such as a chop stick) 11. glasses for mixing acid/base solutions 12. digital camera Procedure 1. Using a sharp knife or razor blade, cut eight uniform red beet root slices, about 3/4" X 1/2" X 1/4" (2 cm X 1.0 cm X 0.5 cm). Try to make the slices as uniform in size as possible. 2. Place the slices in a kitchen glass and run cold tap water gently over them for about 10 minutes to remove betanin released by cells damaged during the sectioning procedure. 3. Using your pH test strips, medicine droppers, vinegar and baking soda solutions, and water, make up a series of different pH solutions: pH 3, pH 7, and pH 9. Be sure not to mix up your droppers and to rinse them with water after each use. It is also important to set up some sort of labeling system for your solutions and test tubes. Transparent tape written over with magic marker works well. 4. Place one section of beet into each of the four test tubes labeled by pH (3, 7, and 9). Add 10 ml (use calibrated markings etched into the side of the test tube to measure) of the corresponding pH solution. 5. Label four test tubes 1D, 2D, 3D, and 4D. Add 10 ml water to each (check that pH ~7) and then add using your medicine dropper, add 1 drop, 2 drops, 3 drops, and 4 drops of detergent to the correspondingly labeled test tube. Using the glass stir rod (or similar thin straight utensil), stir very gently. Be careful not to create bubbles. Gently place one section of beet into each of the four detergent containing test tubes. 6. Incubate all of the tubes at room temperature for at least 30 minutes. 7. Gently shake each tube to equilibrate the released pigment in each one and compare the intensity of color. Record your observations in the lab worksheet results section, and take a digital photo of your results. (Note: Clean out and save your test tubes, you will need them in later experiments.) 8. Complete Part B of the lab#1 worksheet, and move on to Part C. Part C. Wet Lab: Examining Differential Permeability and Osmosis Introduction In this exercise you will use dialysis tubing to simulate differential permeability of cell membranes. Dialysis tubing is a synthetic material that contains pores of a determined size. If a molecule placed in the tubing is larger than the pore size, it will not leak out. You will make up a starch solution and place it in a sac made up of dialysis tubing, which you will place in a glass containing an iodine solution. Iodine in water is a pale yellow-amber color. In the presence of starch, it turns a purple/black color. Thus, you can use the color of iodine to follow the movement of iodine and starch in and out of the dialysis bag. Materials 1. dialysis tubing (3.5”X12”)
X19A - Module 2 2. heat proof, clear glass container 3. 0.5 g cornstarch (¼ teaspoon) 4. 250 ml boiling water (~1 cup) 5. one test tube 6. iodine 7. water 8. a kitchen glass 9. medicine dropper 10. small rubber band 11. water proof marking pen 12. strong tape (duct tape) 13. ruler 14. digital camera Procedure Make the starch solution as follows: 1. Place 250 ml (~ 1 cup) of water in a saucepan on the stove top and bring the water to a gentle boil. 2. Slowly add ~ 0.5 grams (¼ teaspoon) starch into the water & stir to create a cloudy slurry. (be careful not to burn your-self from the steam. Use a long wooden or plastic spoon to stir until the starch is completely dissolved in the water. 3. The solution should be clear , not cloudy. 4. Allow the solution to slowly cool to room temperature. Make sure your starch solution is sufficiently dissolved. 5. Do quick test of the starch by placing a drop of iodine on a paper plate and adding one drop of starch to the iodine drop. It should turn a deep blue/purple color. Prepare the dialysis bag as follows: a. Soak the tubing (~in a glass of water until it is soft and pliable. b. Gently tie a tight knot at one end of the tube. 1. Prepare a solution of iodine in a large kitchen glass by combining water and just enough iodine to form a light-yellow solution. Set aside. 2. Gently tease open the other end of the dialysis tubing and use the medicine dropper to fill it 1/3 of the way (~4 inches) with the starch solution. Using a rubber band, tightly secure the top of the starch-filled dialysis sac. 3. To be sure no starch solution is on the outside of the dialysis membrane, gently rinse off the outside of the bag by placing it briefly under lightly running water. 4. Hold the dialysis tube next to the ruler and record the height of the starch solution that is inside the bag. Take a digital photo. 5. With tape, secure the top of the bag to the outside lip of the large kitchen glass containing the prepared iodine solution. Immerse the starch-filled portion of the bag into the iodine solution.
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