1. Mutagenesis and Cancer
Kamena Kostova and Darienne Myers

2. Overview of Darienne and Kamena’s Lessons
4 Lessons total on Cancer and Mutagenesis
1. Introduction to Cancer and Mutagenesis (Thursday 3/6)
2. Experiment Day! Testing Mutagenesis with Yeast (Friday 3/7)
3. Data Analysis – Is the compound you tested a mutagen?? (Monday, 3/10)
4. Second Experiment Day! Make your compound more or less mutagenic (Thursday 3/13)
5. Group Presentations to discuss your experiments and results (next Friday 3/21)

3. Yeast food If the yeast don’t have the URA gene, 5-FOA is NOT toxic.
Plate with 5-FOA
Yeast
(have the URA gene)
Yeast DIE ☹
5-FOA is a chemical that is toxic to yeast if the yeast have the URA gene.
If the yeast don’t have the URA gene, 5-FOA is NOT toxic.

4. Yeast food
Yeast
(mutation in
the URA gene)
Plate with 5-FOA
Yeast GROW! ☺
5-FOA is a chemical that is toxic to yeast if the yeast have the URA gene.
If the yeast have a mutation in the URA gene, 5-FOA is NOT toxic.

5. Mutagenesis experiment
Plate with 5-FOA
Yeast DIE ☹ Y
Plate with 5-FOA
Yeast GROW! ☺
Yeast
(have the
URA gene)
Yeast
(have a mutation
in the URA gene)
On Friday, we started with yeast that HAVE the URA gene. Do they grow on 5-FOA plates?
NO. They do not grow.

6. Mutagenesis experiment
Plate with 5-FOA
Yeast DIE ☹ Y
Plate with 5-FOA
Yeast GROW! ☺
Yeast
(have the
URA gene)
Yeast
(have a mutation
in the URA gene)
What do you think will happen if we add mutagens to the yeast and grow on 5-FOA?
Treat with
mutagens

7. Mutagenesis experiment
Plate with 5-FOA
Yeast DIE ☹ Y
Plate with 5-FOA
Yeast GROW! ☺
Yeast
(have the
URA gene)
Yeast
(have a mutation
in the URA gene)
What do you think will happen if we add mutagens to the yeast and grow on 5-FOA?
Treat with
mutagens
Some yeast
will get
mutations
in the URA gene
Plate with 5-FOA
Yeast with mutations
GROW
Yeast without mutations
DIE
Do this interactively with the students. “Which yeast will grow on the normal plate?” “Which yeast will grow on the 5-FOA plate?”. Then can go through the cartoon answers on the next slide.

8. Mutagenesis experiment
Plate with 5-FOA
Yeast DIE ☹ Y
Plate with 5-FOA
Yeast GROW! ☺
Yeast
(have the
URA gene)
Yeast
(have a mutation
in the URA gene)
What do you think will happen if we add mutagens to the yeast and grow on 5-FOA?
Treat with
mutagens
Some yeast
will get
mutations
in the URA gene
Yeast with mutations GROW
Yeast without mutations DIE
Plate with 5-FOA
Do this interactively with the students. “Which yeast will grow on the normal plate?” “Which yeast will grow on the 5-FOA plate?”. Then can go through the cartoon answers on the next slide.

9. Mutagenesis experiment
Plate with 5-FOA
Yeast DIE ☹ Y
Plate with 5-FOA
Yeast GROW! ☺
Yeast
(have the
URA gene)
Yeast
(have a mutation
in the URA gene)
What do you think will happen if we add something that is NOT a mutagen to the yeast and grow on 5-FOA?
Treat with a non-mutagen
Yeast will still have the URA gene
Plate with 5-FOA
Yeast without mutations
DIE
Reinforce that the yeast we are working with aren’t actually blue and green ;)

10. Our Mutagenesis Experiment
What did we do last week?
We took yeast and mixed it with water (control) or a potential mutagen (shampoo, makeup, perfume, hot sauce, cheetos, etc) and put them onto plates.
Do you expect to see any colonies for your CONTROL (water)? No
Do you expect to see colonies if what you tested IS a mutagen?
Yes
Do you expect to see colonies if what you tested IS NOT a mutagen?

11. Kamena, Darienne, and Mr. Cohbra’s Experiment
Control
Why is there a colony on the control??
Some possible reasons:
-human error
-some mutations occur randomly
-cell stress can lead to mutations
Ask: what looks strange here? See if they bring up that we do not expect to see a colony on the control.
Ask for some suggestions about why there might be a colony there

12. Do you think it is a mutagen?
What if we see 0 colonies on the control plate and 20 on the experimental plate?
Ask to answer yes or no (or could hold up the blue and white cards again?)

13. Do you think it is a mutagen?
What if we see 0 colonies on the control plate and 20 on the experimental plate?
What if we see 1 colony on the control and 20 on the experimental?
Ask to answer yes or no (or could hold up the blue and white cards again?)

14. Do you think it is a mutagen?
What if we see 0 colonies on the control plate and 20 on the experimental plate?
What if we see 1 colony on the control and 20 on the experimental?
What if we see 19 on the control and 20 on the experimental?
Ask to answer yes or no (or could hold up the blue and white cards again?)

15. Do you think it is a mutagen?
What if we see 0 colonies on the control plate and 20 on the experimental plate?
What if we see 1 colony on the control and 20 on the experimental?
What if we see 19 on the control and 20 on the experimental?
Are the differences we observe because the substance is a mutagen, or just because of random chance?
Ask to answer yes or no (or could hold up the blue and white cards again?)

16. Do you think it is a mutagen?
What if we see 0 colonies on the control plate and 20 on the experimental plate?
What if we see 1 colony on the control and 20 on the experimental?
What if we see 19 on the control and 20 on the experimental?
Are the differences we observe because the substance is a mutagen, or just because of random chance?
Scientists use statistics (math!) to help answer this question
Ask to answer yes or no (or could hold up the blue and white cards again?)

17. Statistical Tests
Statistics tell scientists whether a difference between their control and their test is because of the experiment they did or because of random chance
The test we’ll use is called Chi-Square

18. Statistical Tests - Chi Square
Chi square test tells us if our observations happen by chance or if there is some interesting biology behind them
VS.

19. Statistical Tests - Chi Square
Chi square test tells us if our observations happen by chance or if there is some interesting biology behind them
We compare our results to what we expect to see if everything was happening by chance!
The idea that our observations are due to randomness is called null hypothesis.
In our case, the null hypothesis is that the substance tested is not a mutagen and any difference between the control and the tested substance is by chance.

20. Statistical Tests - Chi Square
We’ll use the following formula to calculate chi square

21. Statistical Tests - Chi Square
Once we obtain a number for chi square, we will compare it to a reference value, 3.84 in our case.
-> if chi square > 3.84 => the difference we see is due to the experimental conditions (the substance IS a mutagen)
-> if chi square < 3.84 => the difference is due to random chance (the substance is NOT a mutagen)

22. Statistical Tests: Coin Flipping
Make sure everyone has a coin. Explain the idea of a fair and biased coin

23. Statistical Tests: Coin Flipping
Do you think the coin you are going to flip is fair, or is it biased towards heads or tails?
Write down the null hypothesis
Go through the first section of the worksheet as a group. Then we can pass out coins or they can get them out, they can do the 20 flips, and then we can gather as a group and have 1-2 groups share their results.

24. Statistical Tests: Coin Flipping
Do you think the coin you are going to flip is fair, or is it biased towards heads or tails?
Write down the null hypothesis
The expected outcome is that it is equally likely to get heads and tails when you flip the coin.
So, if you flip the coin 20 times, how many times will you get heads? How many tails?
Go through the first section of the worksheet as a group. Then we can pass out coins or they can get them out, they can do the 20 flips, and then we can gather as a group and have 1-2 groups share their results.

25. Statistical Tests: Coin Flipping
Do you think the coin you are going to flip is fair, or is it biased towards heads or tails?
Write down the null hypothesis
The expected outcome is that it is equally likely to get heads and tails when you flip the coin.
So, if you flip the coin 20 times, how many times will you get heads? How many tails?
10 heads, 10 tails
Go through the first section of the worksheet as a group. Then we can pass out coins or they can get them out, they can do the 20 flips, and then we can gather as a group and have 1-2 groups share their results.

26. Statistical Tests: Colony Counting
Do you think that the substance you’ve tested is a mutagen?
Write down the null hypothesis
Go through the first section of the worksheet as a group. Then we can pass out coins or they can get them out, they can do the 20 flips, and then we can gather as a group and have 1-2 groups share their results.

27. Statistical Tests: Colony Counting
Do you think that the substance you’ve tested is a mutagen?
Write down the null hypothesis
The expected outcome is that the yeast are equally likely to grow when treated with water or your substance.
So, if you’ve counted 20 colonies total on your plate, how many colonies do you expect to see on the control third of the plate? How many colonies on the test third of the plate?
Go through the first section of the worksheet as a group. Then we can pass out coins or they can get them out, they can do the 20 flips, and then we can gather as a group and have 1-2 groups share their results.

28. Statistical Tests: Colony Counting
Do you think that the substance you’ve tested is a mutagen?
Write down the null hypothesis
The expected outcome is that the yeast are equally likely to grow when treated with water or your substance.
So, if you’ve counted 20 colonies total on your plate, how many colonies do you expect to see on the control third of the plate? How many colonies on the test third of the plate?
10 on the control, 10 on the test
Go through the first section of the worksheet as a group. Then we can pass out coins or they can get them out, they can do the 20 flips, and then we can gather as a group and have 1-2 groups share their results.