Cell Division Lab Austin Cao, AP Biology Part A: Getting started Mitosis is the process by which a cells nucleus

replicates itself. This is an integral part of the cell cycle that allows cells to quickly reproduce themselves. Meiosis is the process by which a cell splits its number of chromosomes in half into gametes used for sexual reproduction. 1. How did you develop from a single-celled zygote to an organism with trillions of cells? My cells simply underwent mitosis many many many times. 1+2+4+8+16... sum = 100 trillion. 2. How is cell division important to a single-celled organism? Cell division allows the organism to asexually reproduce. 3. What must happen to ensure successful cell division? DNA must be duplicated successfully, and all checkpoints must be passed. 4. How does the genetic information in one of your body cells compare to that found in other cells? They are identical. 5. What are some advantages to asexual reproduction in plants? No partner is needed for reproduction. Genetic code does not vary between generations. 6. Why is it important for DNA to be replicated prior to cell division? The amount of DNA must be constant from generation to generation. 7. How do chromosomes move inside a cell during cell division? Mitotic spindle fibers guide them. 8. How is the cell cycle controlled? What would happen if the control were defective? A checkpoint system controls the cycle. Defective control may allow rampant cell division. Part B: Modeling mitosis Prophase: Chromosomes form, nucleus dissolves, DNA has replicated.

Metaphase: Spindle fibers from centrosomes attach to chromosomes.

Anaphase: Chromosomes are pulled apart to edges of cell.

Telophase and Cytokinesis: Cleavage furrow begins to form.

Interphase: Two daughter cells are formed.

Chromosome 1 1 2 3 4 5 6 g/g g/g g/g g/g g/c g/c

Chromosome 2 r/r r/r r/b r/b r/r r/r

Chromosome 3 b/b b/r b/b b/r b/b b/r

1. If a cell contains a set of duplicated chromosomes, does it contain any more genetic information than the cell before the chromosomes were duplicated? It contains twice as much genetic information. It just two copies. 2. What is the significance of the fact that chromosomes condense before they are moved? This ensures that DNA strands dont break during mitosis/meiosis. 3. How are the chromosome copies, called sister chromatids, separated from each other? Spindle fibers pull them apart. 4. What would happen if the sister chromatids failed to separate? One cell will have two chromatids, the other will have none of this chromosome (some mutation).

Part C: Chi-square analysis

Indiv. Observed Expected Dev (O E) (O-E) 2 (O-E) 2/E X2 Brown 219 150 69 4761 31.74 122.69 Blue 209 288 -79 6241 21.67 Orange 185 289 -104 10816 37.43 Green 186 138 48 2304 16.70 Red 173 139 34 1156 8.32 Yellow 182 150 32 1024 6.83 Total 1154 1154

0 0

Class Observed Expected Dev (O E) (O-E) 2 (O-E) 2/E X2

Brown 201.455 136.76 64.695 4185.44 30.60 66.48

Blue 179.27 263 -83.73 7010.71 26.66

Orange 194.182 263 -70.818 5015.19 19.07

Green 156.455 126.24 30.215 912.95 7.23

Red 161.545 126.24 35.305 1246.44 9.87

Yellow 159.09 136.76 22.33 498.63 3.65

Total 1052 1052

0 0

1. What is a null hypothesis? It our default position: that there is no significance/relationship for what we are investigating. 2. What was our null hypothesis for the M&M Lab? All of the colors had the same frequency. 3. Based on our sample, should we accept or reject our null hypothesis? Reject the null. 4. If we did reject our null hypothesis, what might be some possible explanations for this outcome? Mars thinks that we prefer certain colors, so they alter the frequencies. 5. If we did accept our null hypothesis, what might be some possible explanations for this outcome? Mars just doesnt care about our preferences. Part D: Effects of environment on mitosis 1. What is your experimental hypothesis? Your null hypothesis? Are these the same? Experimental is that lectin decreases rate of mitosis in root cells. Null is it does not affect it. No. 2. How would you design an experiment with onion bulbs? Grow 2 sets of bulbs in separate environments, one treated with lectin. Measure growth of the bulb. 3. What would you measure, and how would you measure it?

You should measure mass with a weight. 4. What would be an appropriate control for your experiment? Grow the bulb in normal conditions. Indiv. Control Lectin Total Interphase 13 25 38 Mitotic 2 2 4 Total 15 27 42

Class Control Lectin Total

Interphase 43 50 93

Mitotic 236 135 371

Total 279 185 464

2. Find your expected values: 185*(43/279)=28.5, 185*(236/279)=156.5 3. Calculate the chi-square value for the test: ((50-29)^2)/29=15.2 ((135-157)^2)/157=3.1 4. Compare this value to the critical value in Table 2. Way greater than 3.84. Reject the null hypothesis.

X^2=18.3

What does it mean if your null hypothesis is rejected and was yours rejected or not? We rejected it, meaning that lectin affects mitosis in root cells. 1. What was the importance of collecting the class data? Class data gives more trails, and thus a more accurate observed data. 2. Was there a significant difference between the groups? Yes there was. 3. Did the fungal pathogen lectin increase the number of root tip cells in mitosis? No it decreased the number of cells in mitosis. 4. What other experiments should you perform to verify your findings? Repeating the experiment in other plants. Is there a more accurate way to count cells in mitosis? 5. Does an increased number of cells in mitosis mean that these cells are dividing faster? Yes, a freeze-frame of more cells in mitosis indicates that they are dividing faster on average. 6. What other way could you determine how fast the rate of mitosis is occurring in root tips? You could also mass the roots over time. Part E: Design your own investigation

Objective: To determine the effect of various luminosities of light on the rate of mitosis of root tips. Procedure: 1. Grow onions in environments of 3 different light conditions. Say, 10 onions each treatment. 2. Cut off the root tips and prepare slides of each onion root tip. 3. Count the mitotic cell percentage. 4. Compare to the control light treatment. My hypothesis is that less light means less energy means less mitosis will occur. Part F: Cancer cells 1. How is the cell cycle controlled in normal cells? The cell cycle is controlled by checkpoints. Cyclins activate respective enzymes to progress the cell. 2. What are cyclins and cyclin-dependent kinases? What do these proteins do in a cell? Cyclins are proteins that bind to cyclin-dependent kinases to activate them, and regulate the cycle. 3. How are normal cells and cancer cells different from each other? Cancer cells continue to divide, uninhibited by factors that normally slow down normal cell growth. 4. What are the main causes of cancer? Any genetic mutation (caused by heritage, radiation, chemicals, etc.). 5. What goes wrong during the cell cycle in cancer cells? The cell passes through checkpoints unchecked despite having damaged DNA. 6. What makes some genes responsible for an increased risk of certain cancers? Some genes code for the checkpoints, so potential mutations may promote cell division. 7. Do you think that the chromosomes might be different between normal and cancer cells? There will be small differences in crucial parts of the DNA. Case 1 1. What went wrong in Henrietta Lackss cervical cancer cells that made them cancerous? A mutation in her DNA- specifically something that gave HeLa cells telomerase, preventing the shortening of telomeres in cell death. 2. How does infections with HPV increase the risk of cervical cancer? HPV can cause DNA mutations that cause cancer. Im not sure how that specifically happens. 1. Should tissue be removed from a patient without his or her consent for research? No. People have rights. 2. How was the HeLa cell line cultured? The cells are placed under controlled conditions, in a specific medium, so that they can grow. 3. What virus infected Henrietta Lacks and may have caused her cervical cancer? What cellular process is affected by this virus? HPV infected her, it affects mitosis (explained above). 7. Do companies or universities have the right to patent discoveries made using her cells? Yes. Who else could you possibly give the patent to? Case 2 1. What happens in a normal cell if the DNA has mutations? The regulation system blocks the cell at a checkpoint, and it undergoes apoptosis. 2. What would happen if cells with mutated DNA replicated?

The mutation would be in the daughter cells. 3. How do cells monitor DNA integrity? The checkpoints. Enzymes search for strand breaks. 4. How are the chromosomes different in the cancer cells compared to normal cells? The chromosomes have some specific mutation in an important locus. 5. How could these differences lead to cancer? Different DNA will code for certain defects, specifically unregulated cells grow into cancer tumors. Part G: Karyotypes Chromosome set #2, Gender: Female, Abnormal or Normal: Abnormal Chromosomal Abnormality: translocation in #9, deletion in #22, Myeloid Leukemia Part H: Modeling meiosis Interphase: Chromosomes formed, nucleus dissolves.

Prophase 1: Chromosomes are duplicated.

Prophase 1: Chromosomes align.

Prophase 1: Homologous chromosomes cross over.

Metaphase 1: Spindle fibers attach.

Anaphase 1: Chromosomes separate.

Cytokinesis 1: Daughter cells are formed.

Metaphase 2: Chromosomes align.

Anaphase 2: Chromatids separate.

Cytokinesis: Cleavage forms.

Four daughter cells are formed.

1. How do sexually reproducing organisms produce gametes from diploid progenitors? Meiosis splits the number of chromosomes (from diploid to haploid). 2. How does the process increase gamete diversity? Crossing over and independent assortment allow for many variations (from father or mother). 3. What are the outcomes from independent assortment and crossing over? Independent assortment results in 2 different gametes. Crossing over results in infinitely many. 4. How does the distance between two genes affect crossover frequencies? The further away they are, the more likely for crossover to occur. 1. When is the DNA replicated during meiosis? Interphase. 2. Are homologous pairs of chromosomes exact copies of each other? No. Crossing over would make no sense then. 3. What is crossing over? Homologous chromosomes randomly exchange genes. 4. What physical constraints control crossover frequencies? Locus on the chromosome. 5. What is meant by independent assortment? When chromatids split apart, they can go in either gamete. Allows for more genetic variation. 6. How can you calculate the possible number of different kinds of gametes? Multiply all of the independently assorted possibilities, and all of the crossovers... impossible. 7. What happens if a homologous pair of chromosomes fails to separate? One cell has a duplicate, one cell has none. This results in diseases like Down syndrome. 8. How are mitosis and meiosis fundamentally different? Meiosis halves the number of chromosomes, mitosis is perfect replication. Part I: Sordaria Sordaria is a spore-producing fugus. The spore chains are visible. During meiosis, crossing-over will changing the colors of the spores, so we can observed crossing over indirectly with Sordaria.

#of Asci (4:4) 73

# of Asci (crossover) 27

Total # of Asci 99

% Asci (crossover)/2 13.5

1. How do you explain the differences between the recombinant asci and the parental types? Im guessing that crossovers are less likely than normals. Parental types are like their parents. 2. What meiotic event can account for this difference? Crossover. 1. Why did you divide the percentage of asci showing crossover (recombinant) by 2? Only half are the result of crossover. 2. The published map distance is 26 map units. How did the class data compare with this distance? 13.5 isnt really close at all? 3. How can you account for any disparities between the class data and the published data? Student error. Or maybe it was intended? 4. Illustrate what happened during meiosis to produce the results you found. Asci branches crossed over with the other asci branches to produce different patterns. 5. Do you think the Philadelphia chromosome is a result of crossing over? No, its a result of failed crossing over. A translocation occurs. 6. Do you think the cell cycle described for mitosis could be applied to meiosis as well? No, they are completely different processes. Part J: Go from here? 1. Can the same (or any) environmental factors you tested above affect the amount of crossing over that occurs in Sordaria? How would you set up an experiment to test this? Grow Sordaria in different conditions and then count the crossovers as above. 2. Revisit the learning objectives stated earlier. Do you better understand mitosis and meiosis? Could you teach this to another class? Yes. Its a pretty tangible, mechanical concept. 3. How do the mechanisms of cell replication affect genetic diversity and evolution? Crossing over, independent assortment, segregation, nondisjunction, and random fertilization all are mechanisms increase genetic diversity. This allows evolution to progress faster. 5. Investigate how growth factors affect the cell cycle. This will help you review cell communication. Growth factors act as signalling molecules in the cell cycle. They stimulate cell growth. 6. Research what tumor suppressors do in the cell cycle and which types of cancers may be caused by mutations in tumor suppressor genes. BRCA2 is a tumor suppressor gene on chromosome 13. It encodes for a protein that fixes mistakes in DNA code. 100s of mutations in BRCA2 can increase the likelihood of breast cancer.