Part C: UV Experiments

• Clouds and haze in the lower atmosphere (troposphere) remove some UV photons.
• The angle of the sun. When the sun is directly overhead (zenith angle = 0o), the most UV gets through. When the sun is lower in the sky--in the winter, early or late in any day, and in places that are far from the equator--there is less UV.
• Higher altitudes, not because there is less ozone but because there is less haze.
• Shading the skin with clothing or anything else cuts off direct exposure to the UV. 
• Sunscreen preparations containing chemicals that absorb UV photons.
• Glass and most plastics absorb UV photons, some plastics more than others. UV absorbing films can be put on the surface of glass and plastics.
• The skin pigment, melanin, absorbs UV and helps protect dark-skinned people.

  The cells of all organisms have ways of repairing most of the DNA damage caused by UV photons. But occasionally the cell's repair machinery makes a mistake which results in a mutation, an error in one of the cell's genes. Many scientists believe that mutations in skin cells sometime cause skin cancer. While sunburn is an immediate effect of UV damage, mutations that may lead to skin cancer accumulate over a long period of time. Experiment: Ordinary yeast cells cannot measure the UV damage because they repair their DNA efficiently. Dr. John Game has developed a UV sensitive strain of yeast by introducing mutations in genes necessary for the repair of DNA. You can use these UV sensitive cells to measure the killing effect of sunlight on these cells. Time Line:
  1st Day: 15 min. Getting Ready
  2nd Day: 45 min. Making the UV Exposure Plates
  3rd Day: 10 min. Observe the growth
  4th Day: 30 min. Observe the growth and discuss the results

  Materials:

  Materials for each student:
  

  • 1 Petri plate containing YED agar medium 1 Alcohol wipe
  • 1 mL sterile calibrated bulbed transfer pipet

  Common Materials:
  Freshly prepared suspension of G948-1C/U yeast cells in water in a sterile specimen jar or in a 50 mL beaker, sterile water Sterile toothpicks (new toothpicks are sterile until the box is opened.) Heavy paper, dark tape or other light- proof material to shade 1/2 of plate, Sunscreen Glass microscope slide Sunglasses, or other material to be tested for their ability to absorb UV Tape Personal Protective Equipment:
  

  • Sun hat, sunscreen, sunglasses.
  
  Getting Ready:
  Time Line: 1st Day:15 min.

  To have a freshly-grown culture of yeast cells, you must grow the G948-1C/U strain on YED growth medium overnight at 30o C or 1 to 2 days at room temperature.
  1. Make a clean sterile work space by wiping the table or bench with an alcohol wipe.
  Select a work place free from drafts since most contamination is airborne.
  2. Open the yeast storage vial.
  Use the broad end of a sterile toothpick to take a small amount of yeast from the slant.
  Replace the lid. Tighten.
  3. Open the YED Petri dish just enough so that you can reach into it with the toothpick full of cells.
  Gently make several streaks of the culture on the surface of the agar. (Remember that you need not be able to see the streaks to have enough to grow into a visible culture overnight.) Close the lid.
  4. Incubate the culture overnight at 30oC, or 2 days at room temperature. (Most microbial cultures should be incubated with the agar side up to prevent condensation from dropping on the colonies.)
  Teacher tip
  

  
                        Mid am     Noon     Mid pm
  
  Summer                3-4        2-3       3-4
  
  Spring &Fall          5-6        3-4       4-5
            
  
  Winter                40-50      15-20     20-30
  
  

  Making the UV exposure plates: click here for the cartoon

  Time Line: 2nd Day: 45 min.

  1. Make a visibly turbid suspension: scrape some yeast with a toothpick and wiping them onto the inside of the sterile jar.
  Add about 35 mL of sterile water.
  Replace the lid and swirl to mix.
  If the suspension is not visibly cloudy, add more yeast cells. 2. Swirl the jar to resuspend the cells before removing each sample.
  3. Pipet 1 mL of the suspension directly onto the surface of a YED agar plate. Replace the lid of the plate.
  4. Tilt and rotate the plate to spread the cells over the surface of the agar. If the liquid did not cover, use the blunt end of a toothpick to spread the suspension.
  5. Let the agar absorb the liquid until it disappears (about 10 minutes).
  6. Secure the lid to the bottom of the Petri dish using a small piece of tape on the side of the dish. (The plastic lid of the Petri dish does not absorb a significant amount of the UV-B found in sunlight, but the tape will.)
  7. Cover half of the lid with a card, opaque tape, or a light-proof envelope.

  8. Cover part of the exposed lid area with sunscreen or anything else you wish to test for its ability to absorb UV.

  9. Expose the plate to direct sunlight outdoors for the number of minutes given in the table.

  Teacher Tip

  10. Hold the plate so that the surface of the agar is perpendicular to the sun's rays.
  At this position the shadow will be smallest.
  Place a card under the plate to help show the shadow.
  While you are exposing your plate, observe the approximate angle of the sun.
  

  11. Look at your plate each day for three or four days. Make notes on the appearance of your plate every day and answer the questions below.

  Questions:

  1. How does the plate look different the day after it was exposed to the sun (3rd day)?

  2. Describe the plate on 4th day (5th day, if incubated at room temperature).

  3. How can you tell if you gave the exposed half of the plate a lethal dose? Did all of the cells die?

  4. Why did you cover a portion of the plate with a piece of opaque material?

  5. Why is it important to hold the plate perpendicular to the sun's rays?

  6. How can you tell if you have the plate perpendicular to the sun's rays?

  7. How does the angle of the sun affect your chance of getting a sunburn?

  8. How does the angle of the sun affect the amount of UV that falls on the cells in a given amount of time?

  9. How does the time of day and time of year affect the sun's angle and the time required for the cells to receive a lethal exposure?

  10. How does the geographic location affect the time required for the cells to receive a lethal exposure at noon on June 22, when the sky is clear?

  Click here for answers

  Procedure for exposing yeast cells to sunlight. 1) Swirl to suspend fresh yeast in water 2) Pipet suspension onto YED plate 3) Tilt and rotate to spread lawn of cells 4) Cover half of plate 5) Expose to sun
  
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  Last updated Wednesday, 04-Dec-2002 14:59:16 CST