Biology: The Dynamic Science (MindTap Course List)
4th Edition
ISBN: 9781305389892
Author: Peter J. Russell, Paul E. Hertz, Beverly McMillan
Publisher: Cengage Learning
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Chapter 18, Problem 12TYK
Discuss Concepts A
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Explain 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.
The exponential nature of PCR allows spectacular increases in the
abundance of a DNA sequence being amplified. Consider a 10-kbp
DNA sequence in a genome of 1010 base pairs. What fraction of the
genome is represented by this sequence; i.e., what is the fractional
abundance of this sequence in this genome? Calculate the fractional
abundance of this target sequence after 10, 15, and 20 cycles of PCR,
starting with DNA representing the whole genome and assuming
that no other sequences in the genome undergo amplification in the
process.
Look at each PCR component listed below. For each one, determine which steps(s) of the PCR reaction (denaturation, annealing or extension) would be directly affect if that component were missing.
Taq polymerase:
Oligonucleotide primers:
DNA template:
Deoxynucleotides (A, T, G and C):
Imagine that you correctly prepare your PCR reaction mixture, but there is something wrong with the thermal cycler. Describe what would happen if:
The thermal cycler was stuck on 940C:
The thermal cycler cycled between 600C and 720C, but never reached 940C
The thermal cycler cycled between 940C and 720C, but never reached 600
Chapter 18 Solutions
Biology: The Dynamic Science (MindTap Course List)
Ch. 18.1 - What features do restriction enzymes have in...Ch. 18.1 - Prob. 2SBCh. 18.1 - What information and materials are needed to...Ch. 18.2 - What is a transgenic organism?Ch. 18.2 - Prob. 2SBCh. 18.3 - What is a restriction fragment length polymorphism...Ch. 18.3 - Prob. 2SBCh. 18.3 - Prob. 3SBCh. 18 - Prob. 1TYKCh. 18 - Prob. 2TYK
Ch. 18 - Why are antibiotic resistance markers such as ampR...Ch. 18 - After a polymerase chain reaction (PCR), agarose...Ch. 18 - A cDNA and a cloned fragment of genomic DNA share...Ch. 18 - Prob. 6TYKCh. 18 - Which of the following is not true of somatic cell...Ch. 18 - Prob. 8TYKCh. 18 - Prob. 9TYKCh. 18 - Prob. 10TYKCh. 18 - Prob. 11TYKCh. 18 - Discuss Concepts A forensic scientist obtained a...Ch. 18 - 13. Suppose a biotechnology company has developed...Ch. 18 - Prob. 14TYKCh. 18 - You learned in the chapter that an STR locus is a...
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- You are working in a molecular biology laboratory and are having challenges with your PCR. You decide to double-check the PCR protocol programmed into the thermal cycler and discover that the annealing temperature was programmed to be 65 °C instead of 50 °C, as you had intended. What effects would this mistake have on the PCR reaction? Suggest potential solution(s).arrow_forwardExplain why DNA ladders are usually included during gel electrophoresis. One aspect of PCR that can be modified is the annealing temperature. In general, higher annealing temperatures show more specificity towards a single template, whereas lower annealing temperatures show less specificity and may bind to multiple regions throughout the genome. Discuss how using an annealing temperature that is too high or too low might influence the results of a PCR assay (and gel electrophoresis results) such as the one used in this study.arrow_forwardMatch the following terms with their definitions and label each component of the PCR mixture in the diagram (use the letters A-D):I. DNA polymeraseII. PrimersIII. NucleotidesIV. Genomic DNA template A. DNA that contains the target sequence that will be replicated using PCR.B. An enzyme that copies the DNA sequence.C. A mixture of 4 nucleotides (A,G,C, and T) that will be polymerized into the replicated DNA sequence.D. A short DNA sequence that allows the enzyme to bind and initiate polymerization.arrow_forward
- The polymerase chain reaction (PCR) is used by scientists to amplify DNA, particularly when the quantity of DNA is very small, mixed, or contaminated with other organisms. Explain (with words) the how PCR works using a diagram to help illustrate this (show a minimum of three cycles to illustrate your point).arrow_forwardDescribe the possible outcome of a PCR experiment in which (a) one of the primers is inadvertently omitted from the reaction mixture and (b) one of the primers is complementary to several sites in the starting DNA sample.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_forward
- The exponential nature of PCR allows spectacular increases in the abundanceof a DNA sequence being amplified. Consider a 10-kbp DNA sequence in agenome of 1010 base pairs. What fraction of the genome does this sequence represent? That is, what is the fractional abundance of this sequence in this genome?Calculate the fractional abundance of this target sequence after 10, 15, and 20 cycles of PCR, starting with DNA representing the whole genome and assuming that no other sequences in the genome undergo amplification in the process.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_forwardYour amplicon from PCR was subjected to AGE for analysis. You are shown the image of the gel loaded with the following lanes: (A) negative control, (B) size ladder (1, 2, 3, 4, 6, and 10 kb), (C) gDNA extract, (D) PCR amplicon. However, due to mishaps while loading the gel with the samples, you are not sure which lane is which. You are shown a diagram of the obtained gel below. a. Label each lane of the gel. Write only the corresponding letters in the wells above. b. Above each band in the size ladder, write its size (in kb). c. Approximate the size (in kb) of the polystyrenase gene. Write your answer above the band corresponding to the gene. Bonus: If you wish to identify the bacterial species in this scenario, what gene is most commonly and routinely sequenced? Answer: __________________________________arrow_forward
- You want to set up 50 µl total volume of a PCR reaction. You have a microfuge tube of Taq polymerase (5 units/ul), and each PCR reaction requires a final of 10 units of Taq polymerase. You have a microfuge tube of Taq Buffer (5X), and each PCR reaction requires a final of 1X Taq Buffer. How much of Taq polymerase and Taq buffer would you add? Select all that apply 10 ul of Taq buffer 2 ul of Taq polymerase 5 ul of Taq buffer 5 ul of Taq polymerasearrow_forwardPolymerase 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?arrow_forwarda) (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…arrow_forward
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