DNA Fingerprinting Lab Questions

DNA Fingerprinting Lab Questions Pre-Lab Focus Questions: Introduction to DNA Fingerprinting 1. Compare the “backbone” of sugar-phosphate arrangement in the side chains of all three figures. Are there any differences? There are no differences in the backbone of the sugar phosphate arrangement. 2. In the above figure, do all three samples contain the same bases? Describe your observations. Each sample has the same bases. 3. Are the bases paired in an identical manner in all three samples? Describe the pattern of the base pair bonding. Each base pair bonds with the same base. Meaning adenine is always with thymine and cytosine is always with guanine. 4. In your attempt to analyze DNA samples from three different individuals, what conclusions can you make about the similarities and differences of the DNA samples? They all have the same nitrogen bases, but they are not in the same order or pattern. 5. What will you need to compare between these DNA samples to determine if they are identical or not identical? The order and pattern of the nitrogen bases needs to be the same from strand to strand in order to be identical. Lesson 1 Restriction Digests pf DNA Samples 1. How many pieces of DNA would result from this cut? 2 2. Write the base sequence of the DNA fragments on both the left and right side of the “cut”. Left: Right: ATG AATTCTCAATTACCT TACTTAA GAGTTAATGGA 3. What differences are these in the two pieces? The right side contains a much longer string than the left side. And the order is different. 4. DNA fragment size can be expressed as the number of base pairs in the fragment. Indicate the size of the fragments [mention any discrepancy you may detect]. The smaller fragment is ____3____base pairs (bp). What is the length of the longer fragment? 11 base pairs 5. Consider the two samples of DNA shown below [single strands are shown for simplicity]: Sample #1: C A G T G A T C T C G A A T T C G C T A G T A A C G T T Sample #2: T C A T G A A T T C C T G G A A T C A G C A A A T G C A If both samples are treated with a restriction enzyme [recognition sequence GAATTC] then indicate the number of fragments and the size of each fragment from each sample of DNA (will cut between G A). Sample #1 Sample #2 # of fragments: _2___ # of fragments: __2__ List the fragment size in ascending order: largest → smallest (example: 20,14) Sample #1 Sample #2 11 bp 23 bp 17 bp 5 bp Lesson 1 Restriction Digestion of DNA Samples Observation Questions 1. Describe the samples of DNA (physical properties). There were no obvious distinctions in the DNA without any microscope. 2. Is there any observable difference between the samples of DNA? Not that we could see with the naked eye. 3. Describe the appearance of the restriction endonuclease mix. It did not have any visible physical traits. Lesson 1 Restriction Digestion of DNA Samples Review Questions 1. Before you incubate your samples, describe any visible signs of change in the contents of the tubes containing the DNA combined with the restriction enzymes. There was no visible change in the contents. 2. Can you see any evidence to indicate that your samples of DNA were fragmented or altered in any way by the addition of EcoRl/Pstl? Explain. No there doesn't appear to be any evidence 3. In the absence of visible evidence of change, is it still possible that the DNA samples were fragmented? Explain your reasoning. Yes, because DNA is microscopic and it would be hard to see the fragments with the naked eye. (answer the next day-after the restriction digest) 4. After a 24 hour incubation period, are there any visible clues that the restriction enzymes may have in some way changed the DNA in any of the tubes? Explain your reasoning. There are no visible clues because the DNA isnt visible for the eye to see. Lesson 2 Agarose Gel Electrophoresis Review Questions 1. The electrophoresis apparatus creates an electrical field [positive and negative ends of the gel]. DNA molecules are negatively charged. To which pole of the electrophoresis field would you expect DNA to migrate (+ or -)? Explain. The DNA should move from negative to positive. Because the DNA started on the negative side it was then attracted to the positive side. Also the phosphate in the DNA is attracted to the positive side. 2. What color represents the negative pole? Black represented negative. 3. After DNA samples are loaded in wells, they are “forced” to move through the gel matrix. Which size fragment (large vs. small) would you expect to move toward the opposite end of the gel most quickly? Explain. The smallest one because it has less to move meaning it will get to the opposite side faster. 4. Which fragments are expected to travel the shortest distance [remain closest to the well]? Explain. The larger ones because it takes them longer to move. (Stop here after day 2 of lab) Post-Lab Thought Questions 1. What can you assume is contained within each band? The DNA fragments of the suspects. 2. If this were a fingerprinting gel, then how many kinds (samples) of DNA can you assume were placed in each separate well? There is one in each well. 3. What would be a logical explanation as to why there is more than one band of DNA for each of the samples? Since the enzyme cut the DNa into separate pieces, there were multiple bands. 4. What probably caused the DNA to become fragmented? The restriction enzyme was meant to cause the fragmentation of the DNA 5. From the gel drawing on page 35, which of the DNA samples have the same number of restriction sites for the restriction endonuclease used? Write the lane numbers. Lanes 2, 3 and 4 6. From the gel drawing on page 35, which sample has the smallest DNA fragment? Lane 5. 7. Assuming a circular piece of DNA (plasmid) was used as starting material, how many restriction sites were there in lane three? (use gel drawing on page 35) Please note that the starting material was a circular piece of DNA. There are two restriction sites. 8. From the gel drawing on page 35, which DNA samples appear to have been “cut” into the same number and size of fragments? Lanes 2 and 4. 9. Based on your analysis of the example gel drawing on page 35, what is your conclusion about the DNA samples in the photograph? Do any of the samples seem to be from the same source? If so which ones? Describe the evidence that supports your conclusion. DNA samples in lanes 2 and 4 seem to be a match because they are seemingly identical. Post-lab: Analysis of Results • • Make a drawing or take a picture and paste it here of the banding patterns from the DNA electrophoresis gel you made in class. Label the lanes and measure the distance in mm each band is from the beginning well and write it next to the gel. Band CS S1 S2 S3 S4 S5 1 21 28 28 28 25 26 2 26 X X X 28 30 1. Give a written description of your gel (use your senses). It feels squishy and smooth and is blue in color with different lines in it, including a giant crack straight through the middle of the gel, making it quite distinctive from all other gels in the classroom. 2. Compare your gel to 2 other lab groups gel, list the lab groups and describe how your results compared to the other 2 lab groups. Group 3: Same size, with very similar consistency and color. Lacks a massive crack though. Group 6: Same shape and size, and they also had the same results and conclusions. 3. Explain if any of the suspect fragments match the crime scene fragments and how the process of DNA fingerprinting produced these results? Suspect 4’s fragments match the crime scene, which makes it appear as if they are the same DNA.