Essentials of Genetics (9th Edition) - Standalone book
9th Edition
ISBN: 9780134047799
Author: William S. Klug, Michael R. Cummings, Charlotte A. Spencer, Michael A. Palladino
Publisher: PEARSON
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Textbook Question
Chapter 17, Problem 18PDQ
We usually think of enzymes as being most active at around 37°C, yet in PCR the DNA polymerase is subjected to multiple exposures of relatively high temperatures and seems to function appropriately at 70–75°C. What is special about the DNA
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a)
(iii)
(iv)
Polymerase chain reaction (PCR) is a technique that enables multiplication of specific DNA
sample at minute amount to millions or billions of copies at a short time span.
(i)
Figure 1 indicates the chemicals added into the PCR reaction tube prior to the
addition of thermostable DNA polymerase. Do you agree with the list? Justify your
answer.
DNA template
Buffer (containing Tris-HCI,
KCI, Mg2+)
ddNTP
Forward primer
Reverse primer
Figure 1
If TA cloning is planned to be carried out after amplification of the gene, which
thermostable DNA polymerase will you select and the reason for your selection?
Why is the optimal annealing temperature vital in this technique? Explain how this
temperature (too high or too low) will affect the efficiency of this reaction.
If the primers you purchased possessed the following information:
Number of Guanine: 4
Number of Adenine: 3
Number of Thymine: 4
Number of Cytosine: 5
Calculate the melting temperature of this primer and estimate the…
The final amount of each primer required in a PCR reaction is 25 picomol. If the total volume of the PCR reaction is equal to 100 µl and the stock concentration of each primer is equal to 0.0025 mM. Calculate the volume of stock primer that needs to be added in order to ensure a primer amount of 25 picomol.
Polymerase chain reaction (PCR) is a technique that enables multiplication of specific DNA
sample at minute amount to millions or billions of copies at a short time span.
(i)
(ii)
Figure 1 indicates the chemicals added into the PCR reaction tube prior to the
addition of thermostable DNA polymerase. Do you agree with the list? Justify your
answer.
DNA template
Buffer (containing Tris-HCI,
KCI, Mg2+)
ddNTP
Forward primer
Reverse primer
Figure 1
If TA cloning is planned to be carried out after amplification of the gene, which
thermostable DNA polymerase will you select and the reason for your selection?
Chapter 17 Solutions
Essentials of Genetics (9th Edition) - Standalone book
Ch. 17 -
CASE STUDY |Should we worry about recombinant DNA...Ch. 17 - Prob. 2CSCh. 17 - Prob. 3CSCh. 17 -
HOW DO WE KNOW?
1. In this chapter we focused on...Ch. 17 - Prob. 2PDQCh. 17 - What roles do restriction enzymes, vectors, and...Ch. 17 - Prob. 4PDQCh. 17 - Prob. 5PDQCh. 17 - Prob. 6PDQCh. 17 - Prob. 7PDQ
Ch. 17 - List the advantages and disadvantages of using...Ch. 17 - What are the advantages of using a restriction...Ch. 17 - The introduction of genes into plants is a common...Ch. 17 - Prob. 11PDQCh. 17 - Prob. 12PDQCh. 17 - Prob. 13PDQCh. 17 - What advantages do cDNA libraries provide over...Ch. 17 - Prob. 15PDQCh. 17 -
16. List the steps involved in screening a...Ch. 17 -
17. In a typical PCR reaction, describe what is...Ch. 17 -
18. We usually think of enzymes as being most...Ch. 17 - How are dideoxynucleotides (ddNTPs) structurally...Ch. 17 - Prob. 20PDQCh. 17 - One complication of making a transgenic animal is...Ch. 17 -
22. When disrupting a mouse gene by knockout, why...Ch. 17 - Prob. 23PDQ
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- In a typical PCR reaction, describe what is happening in stages occurring at temperature ranges (a) 92–95°C, (b) 45–65°C, and (c) 65–75°C.arrow_forwardPut the steps of one PCR cycle in the correct order: The PCR reaction mixture is heated to about 70 degrees, which is the optimum temperature for the polymerase to build the new strands of DNA, starting at the 3' end of the primer. The PCR reaction mixture is heated to 95 degrees Celsius, which denatures the double stranded template DNA. The PCR reaction mixture is cooled to about 50-55 degrees, which allows the primers to find their complementary site on the template and "anneal" thearrow_forwardMost PCR reactions do not use the more expensive types of DNA polymerase, which have DNA proofreading. How might this be a problem in accurately copying specific DNA sequences of the target gene?arrow_forward
- During PCR amplification in preparation for DNA sequencing, why were there different colors at the 3’ ends of the fragments produced? (What did these four colors represent?)arrow_forwardExplain why a positive control and negative control are included in PCR experiments. Explain the three steps involved in each cycle of polymerase chain reaction.Why is loading dye added to the DNA sample for gel electrophoresis? Explain the function of the following components in a PCR reaction:− Primer, dNTP, MgCl, Taq polymerase, buffer.arrow_forwardIn a typical PCR reaction what is happening in stages occurring at temperature ranges (a) 92–95°C, (b) 45–65°C, and (c) 65–75°C.arrow_forward
- For PCR, what property of DNA does temperature influence? Why do the “denaturation” and “annealing” steps proceed at different temperatures?arrow_forwardIn PCR amplification Why is it important to know the length of the sequence you amplify?arrow_forwardIn DNA isolation techniques, a washing step is always done prior to the final resuspension. What is the purpose of this step? In DNA isolation from blood samples, why does the vial for blood storage contain EDTA? In the preparation for DNA isolation in plants, the plant source is refrigerated and ground prior to extraction. Why is this so? Why are DNA pellets air-dried before resuspension in buffer?arrow_forward
- Our PCR samples already contain loading dye, but sometimes this isn’t the case. If your samples didn’t already contain dye and you wanted to load your PCR sample onto an agarose gel, you’d need to add loading dye to the proper concentration. There is a 6X loading dye available for use; how many µl of this loading dye will you add to 10 µl of your sample so that it is at a 1X working concentration? Show your work.arrow_forwardYou are performing PCR for the first time using some new primers that are 25 nucleotides in length with an estimated melting temperature of 65 0C to amplify a 200 nucleotide gene. After PCR, you run an agarose gel on the samples and observe a faint band at 25 nucleotides. What is the best possible explanation for the results? The primers hybridized to multiple sites on the DNA template. Too much template DNA was added to the PCR mixture. The primers dimerized preventing DNA transcription from occurring. A nuclease was in the solution causing degradation of the DNA.arrow_forwardThe process of PCR essentially revolves around three phases. Briefly describe these phases and the events that occur in them. Take note the temperature on which these phases take place.arrow_forward
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