Lab 9: Molecular Biology

5 Points

Part 1: In this lab, you will learn about the fundamental tools of molecular biology: DNA isolation, gel electrophoresis and PCR. To begin, work through the tutorial on DNA isolation below. While completing this part of the virtual lab, be sure to take notes and complete the questions below. The questions throughout this exercise will be your lab worksheet for this week. However, you will want to take good notes as you will be responsible for the information in the video on the lab exam.

https://learn.genetics.utah.edu/content/labs/extraction/

1.) Why would we want to isolate DNA (what can we use it for)?

2.) Where is DNA located? How much DNA can be found in one cell?

3.) What kind of cells were used in this lab to isolate DNA?

4.) What are the four steps to purify DNA?

5.) Name 6 materials used in the DNA isolation procedure.

6.) What is the function of the lysis solution?

7.) What is the purpose of the salt solution?

8.) How does a centrifuge work?

Part 2: Now that we have the DNA isolated, we can use a thermocycler to perform PCR and make many copies of our region of DNA of interest. Click on the link below to complete the virtual PCR lab. While you work through the exercise, take notes and complete the questions below. This exercise works through the steps of isolating DNA from and then performing PCR using bacteria. When you click on the link below, click “launch interactive” then click on “click to enter the lab” after reading through the intro. For this lab, you only need to work through part 2 titled “PCR amplification”. However, you can look through the other steps as well if you are interested in learning more.

https://www.biointeractive.org/classroom-resources/bacterial-identification-virtual-lab

1.) What does PCR stand for?

2.) How many copies of DNA can PCR produce and how long does it take?

3.) What is the purpose of PCR ?

4.) PCR involves repeating cycles of 3 steps. Describe, in detail and in your own words, what those 3 steps are and what is happening in each step.

Part 3: We have isolated DNA and made billions of copies of a region of interest. The last step is to utilize gel electrophoresis to separate and visualize DNA. The presence of DNA of specific lengths can help identify the perpetrator of a crime, determine who the father of a child is, determine what alleles someone has for a particular gene, etc. Follow the link below to complete the lab simulation. Take notes and complete the questions below while doing the experiment.

https://learn.genetics.utah.edu/content/labs/gel/

1.) What is gel electrophoresis used for?

2.) What makes the DNA move when running a gel?

3.) DNA migrates to the _________________ end of the gel.

4.) True or False: Long strands of DNA migrate farther than short strands of DNA.

5.) Why do we also run a standard/ladder?

6.) True or False: because DNA is negatively charged, it will migrate to the negative pole.

7.) How do we know that the current is running in the gel box?

Part 4: Practice interpreting results

DNA was isolated from 3 people in a family to test for the presence of a disease-causing allele. The disease is the result of a 600 bp long insertion in chromosome 16. If the individual does not have the disease-causing allele (negative), their DNA segment that was amplified using PCR would be 500 bp long. If the individual has the disease-causing allele (positive), their piece of DNA would be 1100 bp long. Remember that everyone will have 2 alleles for this gene (one from mom and one from dad). In order to have this disease, you must inherit the disease-causing allele (positive) from both of your parents = homozygous positive.

To determine the size of the DNA, a ladder was run in the first lane (farthest left lane). The ladder contained the following size segments of DNA: 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1200, 1600, 2000 bp.

Ladder Person 1 Person 2 Person 3

1.) Label the size of each fragment of DNA in the ladder lane in the gel above.
2.) Matching: match each person in this experiment to their correct description for this gene based on the data in the gel by placing the correct letter in the spaces below.
Person 1 _________ a. homozygous recessive
Person 2 _________ b. heterozygous
Person 3 _________ c. homozygous negative
3.) Will any of these 3 individuals have this disease? If so, who will and explain how you know.

4.) Person 2 had only 1 band show up in the gel. What is the size of their band of DNA?