Genetic Analysis: An Integrated Approach (2nd Edition)
2nd Edition
ISBN: 9780321948908
Author: Mark F. Sanders, John L. Bowman
Publisher: PEARSON
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Chapter 16, Problem 9P
Translational fusions between a protein of interest and a reporter protein are used to determine the subcellular location of proteins in vivo. However, fusion to a reporter protein sometimes renders the protein of interest non-functional because the addition of the reporter protein interferes with proper protein folding, enzymatic activity, or protein
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Describe the difference between a transcriptional fusion and a translational reporter gene fusion. What sequences should you include in each? What types of information can be garnered from their analyses?
Please include annotated pictures of the reporter gene expression patterns.
There is a hypothetical gene related to the nervous system of Drosophila. Describe all the methods, steps, and key substances you need to obtain to use the following techniques in experimental design to study the gene:
- In situ hybridization (to find the mRNA)
- Immunohistochemistry (to find the protein)
- CRISPR-Cas9 (for loss of function)
- Expression vector (for gain of function)
Microarrays can be used to determine relative levels of gene expression. In one type of microarray, hybridization of red (experimental) and green (control) labeled cDNAs is proportional to the relative amounts of mRNA in the samples. Red indicates the overexpression of a gene and green indicates the underexpression of a gene in the experimental cells relative to the control cells, yellow indicates equal expression in experimental and control cells, and no color indicates no expression in either experimental or control cells.In one experiment, mRNA from a strain of antibiotic-resistant bacteria (experimental cells) is converted into cDNA and labeled with red fluorescent nucleotides; mRNA from a nonresistant strain of the same bacteria (control cells) is converted into cDNA and labeled with green fluorescent nucleotides. The cDNAs from the resistant and nonresistant cells are mixed and hybridized to a chip containing spots of DNA from genes 1 through 25. The results are shown in the…
Chapter 16 Solutions
Genetic Analysis: An Integrated Approach (2nd Edition)
Ch. 16 - 14.1 What are the advantages and disadvantages of...Ch. 16 - Prob. 2PCh. 16 - 3. Genetic maps and physical maps are both...Ch. 16 - 14.5 What are the advantages and disadvantages of...Ch. 16 - 14.6 You have cloned the mouse ortholog (see...Ch. 16 - 14.13 The CBF genes of Arabidopsis are induced by...Ch. 16 - 14.14 When the S. cerevisiae genome was sequenced,...Ch. 16 - 14.15 Translational fusions between a protein of...Ch. 16 - In enhancer trapping experiments, a minimal...Ch. 16 - 14.19 In Genetic Analysis, we designed a screen to...
Ch. 16 - How would you design a genetic screen to find...Ch. 16 - 14.21 The eyes of Drosophila develop from imaginal...Ch. 16 - 14.22 Given your knowledge of the genetic tools...Ch. 16 - Mutations in the CFTR gene result in cystic...Ch. 16 - Prob. 16PCh. 16 - 14.25 How would you conduct a screen to identify...Ch. 16 - In land plants, there is an alternation of...Ch. 16 - 14.27 The Drosophila evenskipped (eve) gene is...Ch. 16 - Prob. 20PCh. 16 - 14.29 As shown in Figure, mutations in the...
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- A mutant strain of Salmonella bacteria carries a mutation of the rho protein that has fully activity at 37°C but is completely inactivated when the mutant strain is grown at 40°C. a)Speculate about the kind of differences you would expect to see if you compared a broad spectrum of mRNAs from the mutant strain grown at 37°C and the same spectrum of mRNAs from the strain when grown at 40°C. b)Are all the mRNAs affected by the rho protein mutation in the same way? Why or why not?arrow_forwardMicroarrays can be used to determine relative levels of gene expression. In one type of microarray, hybridization of red (experimental) and green (control) labeled cDNAs is proportional to the relative amounts of mRNA in the samples. Red indicates the overexpression of a gene and green indicates the underexpression of a gene in the experimental cells relative to the control cells, yellow indicates equal expression in experimental and control cells, and no color indicates no expression in either experimental or control cells.arrow_forwardIn the bacteriophage T7 system used to express recombinant proteins, the gene of interest is fused to T7 promoter and T7 RNA polymerase is separately cloned into the same cell. What is the main reason this system uses T7 RNA polymerase instead of relying on the bacterial RNA polymerase? To restrict the expression of bacterial protein expression To enhance the amount of recombinant protein expression To enhance the expression of bacterial protein expression To restrict the amount of recombinant protein expression To enable the expression of T7 viral protein expressionarrow_forward
- Could quantitative PCR, which uses a DNA-binding dye, to show how many copies of the target DNA sequence could be used to quantify the amount of mRNA in a cell? Would you expect that a metabolically active tissue such as the liver would show more cDNA copies in such a method, compared to less metabolically active tissues such as skin cells? One reason that the types and amounts of mRNAs are quantified in different tissue types is to compare which genes are activated and which are inactive. It used to be thought that any gene that was transcribed was automatically translated. The discovery of RNA-degrading systems shows that the real situation in cells is more complemented. Do you believe that a larger amount of mRNA of a given type, say for alpha hemoglobin in immature red blood cells is a reliable indicator that more alpha hemoglobin protein will be made in those cells?arrow_forwardThe pre-mRNA transcript and protein made by several mutant genes were examined. The results are given below. Determine where in the gene a likely mutation lies: the promoter region, exon, intron, cap on mRNA, or ribosome binding site. a. normal-length transcript, normal-length nonfunctional protein b. normal-length transcript, no protein made c. normal-length transcript, normal-length mRNA, short nonfunctional protein d. normal-length transcript, longer mRNA, shorter nonfunctional protein e. transcript never madearrow_forwardTo identify the following types of genetic occurrences, would acomputer program use sequence recognition, pattern recognition,or both?A. Whether a segment of Drosophila DNA contains a P element(which is a specific type of transposable element)B. Whether a segment of DNA contains a stop codonC. In a comparison of two DNA segments, whether there is aninversion in one segment compared with the other segmentD. Whether a long segment of bacterial DNA contains one ormore genesarrow_forward
- . Let’s say that you have incredible skill and can isolate the white and red patches of tissue from the Drosophila eyes shown in Figure 12-24 in order to isolate mRNA from each tissue preparation. Using your knowledge of DNA techniques from Chapter 10, design an experiment that would allow you to determine whether RNA is transcribed from the white gene in the red tissue or the whitetissue or both. If you need it, you have access to radioactive white-gene DNAarrow_forwardYou are interested in studying a novel gene that appears to be involved in cancer. There is no information about the function of this gene. What would you do to obtain the cDNA for this gene? How would you express this gene and what expression systems might you utilize to study its function and why? How would determine the subcellular localization of this gene in eukaryotic cells? What are alternative methods in case one doesn't work? How would you purify and determine the 3-dimensional structure of this protein?arrow_forwardA graduate student studying the pathogenic bacteria Acinetobacter baumannii made cDNA from planktonic cells and biofilm cells and performed RNA-Seq on both samples. She aligned her sequencing reads to a locus of the baumannii genome as shown. a. Which genes are on an operon together? Explain which data supports this? b. What is the most expressed transcript from the locus in Planktonic culture? c. Order the transcripts from largest increase in relative expression between biofilm and planktonic cells to the largest decrease in relative expression. d. When this data was compared to microarray transcriptional profiling, the microarray data provided lower expression levels for the most highly expressed transcripts. Why did this occur?arrow_forward
- For a specific type of mutation at a given location in a particular gene, identify whether it will affect the size of the mRNA, the protein, or both. How would the mutant appear on a gel in comparison to the originalarrow_forwardWhen disrupting a mouse gene by knockout, why is it desirable to breed mice until offspring homozygous (-/-) for the knockout target gene are obtained?arrow_forwardExpression of recombinant proteins in yeast is an important tool for biotechnology companies that produce new drugs for human use. In an attempt to get a new gene X expressed in yeast, a researcher has integrated gene X into the yeast genome near a telomere. Will this strategy result in good expression of gene X? Why or why not? Would the outcome of this experiment differ if the experiment had been performed in a yeast line containing mutations in the H3 or H4 histone tails?arrow_forward
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