For MB3006

This is a series of articles that I found useful and/or relevant while evaluating the review article by Thomas Rando. You might also refer to this counter argument published in the same issue of Cell or this proposal of an alternate hypothesis, the Mortal Strand Hypothesis, written by Rando himself in 2013.

Please note that when you are searching for scientific information, you should always try to access the primary research that was done to investigate the question you are addressing. I have provided links to relevant papers below. However, there are also a number of on-line resources from experts that offer clear and concise explanations of these topics, and I’m providing links to a few of these as well. This is NOT license to use un-cited literature or unsupported statements in your own work. Also, please be cognisant of the University’s policy on plagiarism. These links are merely to provide you with alternate learning materials in case the first explanation wasn’t clear.

When searching for scientific literature, I highly recommend that you use Google Scholar, NCBI, or other professional search engines. This will improve the overall quality of your results.

Semi-conservative DNA replication:

A movie showing replication in slow motion, with all the components labelled: https://www.youtube.com/watch?v=27TxKoFU2Nw

Another movie showing replication in 3D, including illustrations of histone and chromatin structures, chromosome segregation, transcription to RNA, and translation to proteins: https://www.youtube.com/watch?v=yqESR7E4b_8

Mutation rates in humans:

Evolution by Douglass Futuyma is available in the UoA library.

An explanation of why mutations occur and why they are problematic from the Nature Education blog (also a great general resource):
http://www.nature.com/scitable/topicpage/dna-replication-and-causes-of-mutation-409

How might you measure mutation rates?
http://jvi.asm.org/content/62/2/558.full.pdf+html
http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0043876

How are stem cells different from their progeny?

Cairns 2002 sites this paper describing how bone marrow stem cells are less able to survive base excision repair and these papers demonstrating that they’re more sensitive to mutagens.

What was the 3H-Td experiment and why was it important?

A retrospective by the original first author, Dr. Karl Lark.
A link to the original Lark article (requires JSTOR login).

Early work measured the distribution of markers across generation using density segregation of radioactively labelled DNA. This is a particularly beautiful and important paper. Check it out!

Rando in particular touts these experiments by Potten et al. (See in particular Figure 6 to see what nuclei that retain the radioactive marker look like early in the experiment and after several rounds of division) as being proof of non-random segregation.

What can we learn by looking across multiple species?

Similar findings were reported in the bacterium Escherichia coli, the filamentous fungus Aspergillus nidulans and the plants Vicia faba and Triticum boeticumWhy did they replicate the results in all these different organisms?

I bring your attention to this paragraph in the above retrospective (emphasis mine):

“We soon discovered that established tissue culture lines (HeLa or CHO) had lost this property (Lark et al., 1966). Had we begun with established cell lines, we would probably have concluded that the non-random segregation we had documented in bacteria was not a property of somatic mammalian cell division, and we would have abandoned the investigation. Instead, we speculated that the polyploid nature of these established cell lines had obscured the non-random segregation of diploid chromosome sets. Although the distinction between cell lines that had acquired immortality and primary cell lines with programmed longevity was known (Hayflick and Moorhead, 1961Hayflick, 1965), we had not considered this distinction as a possible explanation for the difference between a primary mouse line and the HeLa or CHO lines.”

What mechanisms might lead to non-random segregation?

Work by Sherley et al. identifying one possible molecular trigger (pay specific attention to figure 1) for asymmetric segregation.

What is p53? More than just an oncogene!

A paper hypothesizing a mechanism and describing possible experiments to test this model.

How persistent is non-random segregation across generations?

Work by Conboy and Rando using multiple markers shows that the less differentiated cell tends to retain the labelled strand.

How have technological advances impacted our ability to address these questions?

Go back over the various links above and look at the years of publication and the quality of the data. What kinds of data are more convincing? Do you think the correct controls have been used in all cases? Do the authors examine a high number of events or just a few? Do you trust their conclusions? Why?

How can we evaluate scientific literature?

How often is a paper retracted, who falsifies data (please note that this article is itself disputed in the literature), why, and what should we do about it?

An examination of how scientists measure whether a paper is good.

Have you found other resources you think would be interesting to discuss? Leave a note in the comments! 

 

 

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Respiration in action

Project 2: Respiration produces O2 and CO2.

Saccharomyces cerevisiae is such a useful organism because it grows via both respiration and fermentation. During respiration, sugar is converted to ATP, or cellular energy, and CO2 is produced as a byproduct. When Saccharomyces cerevisiae is added to dough, it allows the dough to rise by creating pockets of CO2. What’s more, Saccharomyces cerevisiae is a useful tool for studying human cells because our cells use the same method to produce energy. In this project, we will observe the production of CO2 by Saccharomyces cerevisiae and test different sources of sugar for the capacity to provide cellular energy.

 

Materials:

1 balloon

1 packet of yeast

1 water bottle

Warm water

Potential energy sources: white sugar, brown sugar, barley malt, molasses, honey, agave, stevia , sunflower oil

 

Procedure:

  1. Pour 1/4 packet of yeast to the water bottle.
  2. Add 25 mL warm water.
  3. Add your chosen energy source.
  4. Stretch out the balloon so it is easy to inflate.
  5. Place the balloon over the top of the bottle and observe.
  6. Measure the diameter of the balloon after 5 min, 10 min, and 20 min.
  7. Repeat using different energy sources.

 

Results

What energy sources did you choose and why?

 

Make a table showing the time and the diameter of the balloon for each condition.

 

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Design your own experiment

Project 1, Part 3: Test a hypothesis

You now have several tools to learn about fungi. There are three additional agar plates available for your group. Together, think of a question or questions you would like to address and design an experiment to test them. For example, do you think one of your agents would work better at a different dose? Do you think one of the agents can prevent the growth of your environmental sample? Discuss as a group how you might test these questions, then bring your experimental design to Dr. Ballou and collect the required materials. List the steps of the procedure you followed below and record your results.

 

Materials:

 

 

Procedure:

 

 

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What kills fungi?

Project 1, Part 2: What kills yeast?

diffusion disk assayBaker’s yeast is a common fungal species called Saccharomyces cerevisiae in the phylum Ascomycota. We use it to make bread, but it is also a very useful model for understanding human biology and human disease. In this experiment, we will perform a Diffusion disk assay to identify different materials that can prevent the growth of Saccharomyces cerevisiae. These materials can be fugistatic –meaning they slow or prevent growth but don’t kill Saccharomyces cerevisiae-or fungicidal –meaning they kill Saccharomyces cerevisiae. This type of assay is commonly used to test new drugs and figure out dosage.

Materials:

1 nutrient agar plate
5 diffusion disks
Potential fugicidal or fungistatic agents

Procedure

  1. Place 1 drop of the yeast suspension onto the agar plate.
  2. Use the cell spreader to spread the yeast around the whole plate. Spread out from the center and make sure to cover the edges. Allow the plate to dry for 10 minutes.
  3. Using tweezers, place 5 diffusion disks around the plate. Space them out evenly in a circle.
  4. Select 5 agents to test. When choosing these agents, think about what you need to design a good experiment. What controls should you include?        Possible agents:

i.     Rubbing alcohol

ii.     Hydrogen peroxide

iii.     Iron oxide (Fe2O3), Cobalt carbonate (CoCO3), Copper Carbonate (CuCO3), Sodium Chloride (NaCl)

iv.     OR find something to test from your environment. For example, are there plants that you think might have
fungicidal activity? If so, collect samples, grind them up using a mortar and pestle, and suspend in a small volume of water. Pass the suspension through a sterile filter using a syringe.

5.   For each agent, place 1 drop onto the diffusion disk.
6.   Incubate the plates at room temperature for 48 hours.
7.    Record your results by photographing the plate

Record your results:

What agents did you choose and why?

Which agents prevented growth? Which agents failed?

Bonus: How can you test if the effect is fungistatic or fungicidal?

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