Ch. 15 - Recombinant DNA Technology and Its Applications
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Problem 1
What is CODIS? Describe the four most important features of genetic markers used in this system.
Problem 1
What purpose do the bla and lacZ genes serve in the plasmid vector pUC18?Problem 2
Compare and contrast the terms Paternity Index (PI) and Combined Paternity Index (CPI). How does each contribute to paternity determination?
Problem 2
The human genome is 3×10⁹ bp in length. How would your initial answer change if you knew that the average GC content of the human genome was 40%?Problem 2
The human genome is 3×10⁹ bp in length. How many fragments would be predicted to result from the complete digestion of the human genome with the following enzymes: Sau3A (˘GATC), BamHI (G˘GATCC), EcoRI (G˘AATTC), and NotI (GC˘GGCCGC)?Problem 3
Ligase catalyzes a reaction between the 5′ phosphate and the 3′ hydroxyl groups at the ends of DNA molecules. The enzyme calf intestinal phosphatase catalyzes the removal of the 5′5′ phosphate from DNA molecules. What would be the consequence of treating a cloning vector, before ligation, with calf intestinal phosphatase?Problem 3
What is the exclusion principle? How is it used in forensic genetic analysis and in paternity determination?
Problem 4
What is the statistical principle underlying genetic health risk assessment? Why are these assessments not predictive of disease occurrence?
Problem 4
You have constructed four different libraries: a genomic library made from DNA isolated from human brain tissue, a genomic library made from DNA isolated from human muscle tissue, a human brain cDNA library, and a human muscle cDNA library. Which of these would have the greatest diversity of sequences?Problem 4
You have constructed four different libraries: a genomic library made from DNA isolated from human brain tissue, a genomic library made from DNA isolated from human muscle tissue, a human brain cDNA library, and a human muscle cDNA library. Would the sequences contained in each library be expected to overlap completely, partially, or not at all with the sequences present in each of the other libraries?Problem 5
Explain the meaning of 'identity by descent' in the context of identifying genealogical relationship between individuals. In these analyses, why are segments of chromosomes (haplotypes) rather than individual STRs used to identify genetic relationships?
Problem 5
Using the genomic libraries in Problem 4, you wish to clone the human gene encoding myostatin, which is expressed only in muscle cells. Assuming the human genome is 3x10⁹ and that the average insert size in the genomic libraries is 100 kb, how frequently will a clone representing myostatin be found in the genomic library made from muscle?Problem 5
Using the genomic libraries in Problem 4, you wish to clone the human gene encoding myostatin, which is expressed only in muscle cells. How frequently will a clone representing myostatin be found in the genomic library made from brain?Problem 5
Using the genomic libraries in Problem 4, you wish to clone the human gene encoding myostatin, which is expressed only in muscle cells. How frequently will a clone representing myostatin be found in the cDNA library made from muscle?Problem 5
Using the genomic libraries in Problem 4, you wish to clone the human gene encoding myostatin, which is expressed only in muscle cells. How frequently will a clone representing myostatin be found in the cDNA library made from brain?Problem 6
The human genome is 3×10⁹ bp. You wish to design a primer to amplify a specific gene in the genome. In general, what length of oligonucleotide would be sufficient to amplify a single unique sequence? To simplify your calculation, assume that all bases occur with an equal frequency.Problem 6
A victim of murder is found to have scrapings containing skin cells under several of her fingernails. Genetic analysis confirms that the DNA isolated from these cells came from the same individual and does not match the DNA of the victim. The results shown below are for six CODIS STR markers from the crime scene DNA (from under the victim's fingernails and presumed to be the murderer's), and from three suspects (A, B, and C) who have been detained for questioning about the murder. Do the STR results exclude any of the three suspects? Explain.Problem 6
A victim of murder is found to have scrapings containing skin cells under several of her fingernails. Genetic analysis confirms that the DNA isolated from these cells came from the same individual and does not match the DNA of the victim. The results shown below are for six CODIS STR markers from the crime scene DNA (from under the victim's fingernails and presumed to be the murderer's), and from three suspects (A, B, and C) who have been detained for questioning about the murder. Is there a failure to exclude any of the suspects? Explain.Problem 7
Using animal models of human diseases can lead to insights into the cellular and genetic bases of the diseases. Duchenne muscular dystrophy (DMD) is the consequence of an X-linked recessive allele. How would you make a Drosophila model of DMD?Problem 7
The results shown are from a DNA test for four genes used in a paternity identification case. DNA for the mother (M) and her child (C) are shown along with DNA from two possible fathers, F1 and F2. In the 'C' column, label the DNA bands contributed by the mother with 'M' and the DNA bands contributed by the father with 'F.'Problem 7
The results shown are from a DNA test for four genes used in a paternity identification case. DNA for the mother (M) and her child (C) are shown along with DNA from two possible fathers, F1 and F2. Based on the exclusion principle, is either man excluded as the possible father? Explain.Problem 7
The results shown are from a DNA test for four genes used in a paternity identification case. DNA for the mother (M) and her child (C) are shown along with DNA from two possible fathers, F1 and F2. What can you conclude based on the DNA results available?Problem 7
Using animal models of human diseases can lead to insights into the cellular and genetic bases of the diseases. Duchenne muscular dystrophy (DMD) is the consequence of an X-linked recessive allele. How would you make a mouse model of DMD?Problem 8
Figure E.1 illustrates the results of an electrophoretic analysis of 13 CODIS STR markers on a DNA sample and identifies the alleles for each gene. Table E.2 lists the frequencies for alleles of three of the STRs shown in the figure. Use this information to calculate the frequency of the genotype for STR genes FGA, vWA, and D3S1358 given in Figure E.1.Problem 8
Compare methods for constructing homologous recombinant transgenic mice and yeast.Problem 9
Additional STR allele frequency information can be added to improve the analysis in Problem 8. The frequency of D8S1179₁₂ = 0.12. The frequency of D16S539₁₈ = 0.08 and of D16S539₂₀ = 0.21. Lastly, D18S51₁₉ = 0.13 and D18S51₂₀ = 0.10. Combine the allele frequency information for these three STR genes with the information used in Problem 8 to calculate the frequency of the genotype for six of the STR genes.Problem 9
Chimeric gene-fusion products can be used for medical or industrial purposes. One idea is to produce biological therapeutics for human medical use in animals from which the products can be easily harvested—in the milk of sheep or cattle, for example. Outline how you would produce human insulin in the milk of sheep.Problem 10
The frequencies of the four alleles contributed to the child by possible father F1 in Problem 7 are 0.18, 0.23, 0.13, and 0.14. Calculate the Combined Paternity Index (CPI) for the four genes in this analysis.Problem 10
The frequencies of the four alleles contributed to the child by possible father F1 in Problem 7 are 0.18, 0.23, 0.13, and 0.14. Make a statement about the possible paternity of F1 based on this analysis.Problem 10
Why are diseases of the blood simpler targets for treatment by gene therapy than are many other genetic diseases?
