Activity 2.1vkkuiohinklnlk lknlilhli PDF

Preview

Summary

ljhlihlihlknk kpihnjikm;kl ihpihiyftcgvjhgvjh uougoulhlhnl kkuggkbkugugkukjj jbluhoh...

Description

Part I: Overview of the lab Overall Objective:The overall objective of this experiment is to find out whether or not each person has either a homozygous or heterozygous gene towards the taste of a certain chemical, specifically the PTC chemical. It will also determine the differences between Main steps 1. Isolate your DNA - Saline solution to extract DNA; clump using centrifuge - Pour out Supernatant: add celex to resuspended cheek cells to expose the cells DNA - Hot bath and centrifuge complete process - This process exposes your DNA to be further worked upon 2. Amplify your DNA by PCR - Next the exposed DNA undergoes PCR procedures to amplify and copy the DNA so that more copies of the gene of interest are available to be targeted in the proceeding steps. - PCR procedure includes introducing the exposed DNA to a PCR primer and loading into a thermal cycler. 3. Digest PCR Products with Restriction Enzyme Haelll - After enough copies have been made the next step is to isolate the gene of interest. - The restriction enzyme Haelll is introduced to the replicated DNA and cuts any base pair differences recognized as the gene of interest (the gene for tasting). 4. Analyze PCR Products by Gel Electrophoresis - After your DNA was tested for the gene of interest in the last step, Gel Electrophoresis reveals the results. - Depending if or how many times Haelll cut your DNA sequence at the gene of interest will determine your genotype. - Gel Electrophoresis sorts the sequences of DNA by segment length. Fewer cuts will be represented with fewer bands on the Gel and vice versa. 5. Determine your PTC Genotype and Phenotype - By interpreting the results illustrated by the Gel Electrophoresis your PTC Genotype is determined. - Compare this to your phenotype: Your phenotype is found by physically testing your response to PTC paper. 2.

3. Reflection Question: After watching the video, consider the techniques used: using a micropipette, using the thermocycler, casting a gel, and completing gel electrophoresis. Describe which of the skills you are most and least comfortable with and why. I am most comfortable with using the micropipette because its easy to get the right amount of liquid that is needed. The hardest part would be completing the gel electrophoresis, simply because you have to get a certain amount of liquid into the slot and it could be difficult to see if you're actually inserting into the right place, also because you have to be gentle with the gel.

How could you practice the skill(s) you are least comfortable with? I could practice by using a small pipette and inserting the liquid into a small precise slot. Part II: With a partner, brainstorm how you can isolate DNA from your own cheek cells. Thinking about where DNA is located in the cell, describe what has to happen for you to isolate cells and release the DNA from these cells. Write down your ideas in your lab notebook and discuss your ideas with the class. - When salt water is gargled and spit it back out, some of the cheek cells become suspended in the salt water, the more vigorously it is gargled, the more cheek cells will collect in the salt water. The washing up of the liquid breaks down the cheek cell membranes, which causes the DNA to be released into the salt water. Because DNA is

not soluble in alcohol, it forms a solid where the alcohol and salt water layers meet. Most other substances from the cheek cells stay dissolved in the salt water layer. The white strings and clumps, the more DNA molecules are clumped together. Observe the DNA sequences of a “nontaster” and a “taster” shown below. NONTASTER TASTER GGCGGGCACT GGCGGCCACT CCGCCCGTGA CCGCCGGTGA In your laboratory journal, describe how the enzyme would cut in a taster and in a nontaster. How many fragments would be produced in each case? Draw or explain how this product would be verified using gel electrophoresis. - A non-taster would not be cut at all; its genotype is G-C where the taster is cut once and at the cut it is different from the non-taster and the genotype is C-G. GGCGGGCACT GGCGG|CCACT CCGCCCGTGA CCGCC|GGTGA Predict what you would see on a gel if you were a heterozygous Tt taster There will be three strands due to the split by being cut giving two, and a split by not being cut giving you one. PLTW LABORATORY JOURNAL Describe the number of bands you would see on the gel if a person was a tt nontaster (homozygous recessive), a TT taster (homozygous dominant), or a Tt taster (heterozygous). Relate what you would expect to see to SNPs and the restriction digest. - On a gel, someone who was homozygous-recessive would appear to be a very faint and light line on the DNA gel electrophoresis. Someone who is heterozygous, would appear to also have a faint and light line, however, not nearly as light and faint as homozygous recessive would be. Someone who is homozygous dominant would be dark and thicker, but would also have a space between their genetic data from the gel electrophoresis. Describe your findings in your laboratory journal. Predict whether or not they will be able to taste PTC. 3 out of the 4 people would not be able to take the PTC because of their homozygous recessive genes.

Conclusion Questions- Explain how the HaeIII enzyme discriminates between the C-G polymorphism in the TAS2R38 gene. - The HaeIII enzyme discriminates between the C-G polymorphism in the TAS2R38, because the HaeIII doesn't cut at C-G it cuts at CC-GG which doesn't happen on the TAS2R38 gene if you're a nontaster. - Using what you know about genetics, SNPs, and the PTC gene, explain why it is possible for a person to be a “weak taster.” - It is possible to be a weak-taster because everyone gets two of each gene, one from each parent, so if one gets a certain gene that can taste PTC and one that

-

-

-

-

-

-

cannot taste it, the gene for tasting the PTC isn't as strong as someone who both genes taste it. Some studies have shown that PTC “tasters” are less likely to become smokers. Why do you think scientists are seeing this correlation? - PTC tasters are less likely to become smokers because PTC has a bitter taste so if cigarettes taste bitter to them because of the gene they'd be less likely to want to smoke. How can the techniques described in this lab be used to test for human disease genes? Would this type of testing work on every disease with a genetic component? - The techniques described in this lab can be used to test for human disease genes because as long as there's a genetic component that causes the disease all you need is a restriction enzyme that can cut the area of the gene that changed so when you run a gel you can see the multiple strands of DNA telling you if they have the disease. What ethical issues are raised by human DNA typing experiments? - There are specific ethical issues that are raised by human DNA experiments. For example, some believe that it can lead to people knowing too much which can lead to anxiety, some alleged that it could even be an invasion of privacy. Lastly, it may not be affordable for all to afford, so only wealthy people could get the advantages of this, whereas more disadvantaged people. Watch this video about Rita, a Clinical Laboratory Scientist. How would Rita use the techniques you practiced in this lab to test for human disease genes? Would this type of testing work on every disease with a genetic component? - The techniques described in this lab can be used to test for human disease genes because as long as there's a genetic component that causes the disease all you need is a restriction enzyme that can cut the area of the gene that changed so when you run a gel you can see the multiple strands of DNA telling you if they have the disease. What ethical issues are raised by human DNA typing experiments? - There are specific ethical issues that are raised by human DNA experiments. For example, some believe that it can lead to people knowing too much which can lead to anxiety, some alleged that it could even be an invasion of privacy. Lastly, it may not be affordable for all to afford, so only wealthy people could get the advantages of this, whereas more disadvantaged people Another student at the college, Bob, returns to the lab for his PTC test results. The gel shows that he is a homozygous recessive nontaster. However, when Mick tastes the PTC paper, he obviously can taste the bitter chemical. Provide a possible explanation for this difference. - PTC gene has two main alleles; one that codes for tasting and the other non-tasting. The allele for tasting determines a trait of tongue papillae that can bind the PTC molecules. Non-tasting allele forms papillae that are unable to bind PTC effectively. Therefore, the combination of these alleles in the genotype of a person determines their phenotype – PTC sensitivity....