DNA extraction

Explain in your own words what extracting DNA from a eukaryotic cell means, and name two simple materials from which cells can be obtained for the lab (for example, cheek cells or a fruit). Why should the material be thoroughly mashed before starting?

In the DNA extraction lab, dish soap, salt and very cold alcohol are used. Explain the role of each reagent: which cellular structures the detergent dissolves, what the salt is used for, and what happens to the DNA when it comes into contact with the alcohol.

Explain why almost any tissue works for extracting DNA, for example a lettuce leaf or a piece of chicken, as long as its cells have a nucleus. What do all those cells have in common?

Dish soap dissolves cell membranes because these are made of a phospholipid bilayer. Explain the analogy with what the detergent does when washing a greasy plate and what then happens to the contents of the cell.

Look at the diagram of the test tube at the end of the DNA extraction lab. Identify what the labels ?1, ?2 and ?3 indicate (the alcohol layer, the aqueous extract with the dissolved DNA, and the DNA strands appearing at the interface), and explain why DNA concentrates precisely in that area.

In the test tube, the DNA goes from being dissolved in the extract to forming a precipitate of strands when the alcohol is added. Explain whether the DNA molecule has changed chemically in this process or only its physical state has changed, and compare it with what happens to salt when it precipitates in water.

The alcohol in the lab must be very cold and should be poured carefully so that it forms a layer on top of the extract without mixing with it. Explain the advantage of the alcohol being cold for the solubility of DNA, and why the two layers should not mix.

DNA is soluble in the salty aqueous extract but insoluble in cold alcohol, while many proteins and membrane remains stay in the liquid. Explain the difference between dissolving and precipitating, and why this change in solubility allows the DNA to be separated from most impurities.

After mashing the material with the extraction mixture, the lab continues by filtering everything. Explain what is retained by the filter, what passes into the beaker, and why it is useful to get rid of the solid part.

List the laboratory equipment needed for the DNA extraction lab (beaker, strainer or filter, test tube, glass rod) and explain what each item is used for.

Although the practical uses kitchen ingredients, it is carried out in a laboratory. Name two safety rules you must follow (for example, do not taste anything and keep the alcohol away from any flame) and explain the reason for each one.

These steps of the DNA extraction lab are out of order: add the very cold alcohol down the side of the tube without shaking; mash the material; prepare the mixture of water, salt, and detergent; filter the mixture; collect the DNA strands with a rod. Put them in order and justify why each step should be in that position.

Design an experiment to find out which fruit, among banana, kiwi, and strawberry, yields the most DNA using the same protocol. State which variables you would keep constant so that the comparison is fair and which one you would measure to compare the results.

You want to extract DNA from a strawberry and an onion using the same protocol. Reason about what adjustments you might need in mashing or in the amount of detergent depending on the hardness of each tissue, and predict from which one you expect to obtain more visible DNA, knowing that the strawberry is octoploid: its cells have eight sets of chromosomes instead of two.

The DNA obtained with the school protocol is not pure: it precipitates along with proteins (histones) and lipids. One team proposes adding fresh pineapple juice (rich in bromelain, a protease enzyme) to the filtrate before adding the alcohol. Another team proposes heating the filtrate to $100\ ^\circ\text{C}$ for $15$ minutes to eliminate protein impurities.

Evaluate both proposals: justify which of the two is appropriate by explaining the action of the protease on the histones, and predict the counterproductive effect that boiling the sample would have on the DNA molecule.

In a forensic investigation, a tiny biological sample is collected (a hair with a root containing barely a few dozen cells). A student claims: "We can use the homemade strawberry protocol: we crush it, add dish soap, salt, and rubbing alcohol, and then read the genes with a magnifying glass." Refute the claim by identifying three insurmountable limitations of this school method and justify which advanced techniques (such as PCR amplification or enzymatic purification) are essential in a real laboratory to analyze the sample.

At the end of the lab with strawberry and kiwi, a student compares both tubes and states: "The DNA of both fruits looks exactly the same (a whitish, filamentous mass). This proves that at the molecular level, DNA of all species is identical and that extraction cannot be used to differentiate between them". Refute their conclusion by distinguishing which part of the DNA molecule is universal to all living things and which part actually contains the information that allows a laboratory to sequence and identify the species.

During a plant DNA extraction lab, three groups obtain failed results:

• Group A: Blended the plant tissue in a blender at maximum power for ten minutes and does not observe strands, only a cloudy liquid without defined filaments.

• Group B: Added 96% alcohol at room temperature ($22\ ^\circ\text{C}$) instead of precooling it to $-20\ ^\circ\text{C}$, and observes hardly any precipitate at the interface.

• Group C: Vigorously stirred the test tube with a glass rod right after pouring the alcohol, and the fibers that were beginning to clump broke apart.

Justify what occurred at a physical or molecular level in each case to explain the failure of the extraction.

A group of students wants to adapt the plant DNA extraction protocol to animal tissue (chicken breast). They propose eliminating two steps: the initial mechanical grinding (because meat is softer than a plant cell wall) and the addition of detergent (because animal cells lack a cell wall).

Critically evaluate their two proposals: justify whether it is correct to dispense with each of these steps based on animal cell organization, and predict what they would obtain if they omitted the detergent.