DNA Tools and Biotechnology

If the template strand has two or more identical nucleotides in a row, their complementary nucleotides will be added one after the other in the same flow step. How are two or more of the same nucleotide (in a row) detected in the flowgram? (See sample on the right.) Write out the sequence of the first 25 nucleotides in the flow-gram above, starting from the left (Ignore the very short lines.)

The restriction site for an enzyme called PvuI is the following sequence:

5' -CGATCG-3'

3' -GCTAGC-5'

Staggered cuts are made between the T and C on each strand. What type of bonds is being cleaved? (See Concept 5.5.)

One strand of DNA molecule has the following sequence:    

5’-CTTGACGATCGTTACCG-3’

Draw the other strand. Will PvuI (see question I) cut this molecule? If so, draw the products.

What are some potential difficulties in using plasmid vectors and bacterial host cells to produce large quantities of proteins from cloned eukaryotic genes?

Compare Figure 20.7 with Figure 16.20. How does replication of DNA ends during PCR proceed without shortening the fragments each time?

Describe the role of complementary base pairing during RT-PCR, DNA microarray analysis, RNA sequencing, and CRISPR-Cas9 editing.

Consider the microarray in Figure 20.12. If a sample from normal tissue is labeled with a green fluorescent dye and a sample from cancerous tissue is labeled red, what color spots would represent genes you would be interested in if you were studying cancer? Explain.

Based on current knowledge, how would you explain the difference in the percentage of tadpoles that developed from the two kinds of donor nuclei in Figure 20.16?

A few companies in China and South Korea provide the service of cloning dogs, using stem cells from their clients’ pets to provide nuclei in procedures like that in Figure 20.17. Should their clients expect the clone to look identical to their original pet? Why or why not? What ethical questions does this bring up?

Based on what you know about muscle differentiation (see Figure 18.18) and genetic engineering, propose the first experiment you might try if you wanted to direct an embryonic stem cell or iPS cell to develop into a muscle cell. 

What is the advantage of using stem cells for gene therapy or gene editing?

List at least three different properties that have been acquired by crop plants via genetic engineering.

As a physician, you have a patient with symptoms that suggest a hepatitis A infection, but you have not been able to detect viral proteins in the blood. Knowing that hepatitis A is an RNA virus, what lab tests could you perform to support your diagnosis? Explain the results that would support your hypothesis. 

In DNA technology, the term vector can refer to

(A) the enzyme that cuts DNA into restrictionfragments.

(B) the sticky end of a DNA fragment.

(C) a SNP marker.

(D) a plasmid used to transfer DNA into a living cell.

Which of the following tools of DNA technology is incorrectly paired with its use? 

(A) electrophoresis—separation of DNA fragments

(B) DNA ligase—cutting DNA, creating sticky ends of restriction

fragments

(C) DNA polymerase—polymerase chain reaction to amplify

sections of DNA

(D) reverse transcriptase—production of cDNA from

mRNA

Plants are more readily manipulated by genetic engineering than are animals because

(A) plant genes do not contain introns.

(B) more vectors are available for transferring recombinant

DNA into plant cells.

(C) a somatic plant cell can often give rise to a complete

plant.

(D) plant cells have larger nuclei.

A paleontologist has recovered a bit of tissue from the 400-year old preserved skin of an extinct dodo (a bird). To compare a specific region of the DNA from a sample with DNA from living birds, which of the following would be most helpful in increasing the amount of dodo DNA available for testing?

(A) SNP analysis

(B) polymerase chain reaction (PCR)

(C) electroporation

(D) gel electrophoresis

DNA technology has many medical applications. Which of the following is not done routinely at present?

(A) production of hormones for treating diabetes and

dwarfism

(B) analysis of gene expression for more informed cancer

treatments

(C) gene editing by the CRISPR-Cas9 system in viable human

embryos to correct genetic diseases

(D) prenatal identification of genetic disease alleles

Which of the following is not true of cDNA produced using human brain tissue as the starting material?

(A) It can be amplified by the polymerase chain reaction. 

(B) It was produced from pre-mRNA using reverse transcriptase. 

(C) It can be labeled and used as a probe to detect genes expressed in the brain.

(D) It lacks the introns of the pre-mRNA.

Expression of a cloned eukaryotic gene in a bacterial cell involves many challenges. The use of mRNA and reverse transcriptase is part of a strategy to solve the problem of

(A) post-transcriptional processing.

(B) post-translational processing.

(C) nucleic acid hybridization.

(D) restriction fragment ligation.

Which of the following sequences in double-stranded DNA is most likely to be recognized as a cutting site for a restriction enzyme?

(A) AAGG

TTCC

(B) GGCC

CCGG

(C) ACCA

TGGT

(D) AAAA

TTTT

Imagine you want to study one of the human crystallins, proteins present in the lens of the eye (see Figure 1.8). To obtain a sufficient amount of the protein of interest, you decide to clone the gene that codes for it. Assume you know the sequence of this gene. Explain how you would go about this.

Looking at Figure 20.15, what does it mean for a SNP to be “linked” to a disease-associated allele? How does this allow the SNP to be used as a genetic marker?(See Concept 15.3.)

You are cloning an aardvark gene, using a bacterial plasmid as a vector. The green diagram shows the plasmid, which contains the restriction site for the enzyme used in Figure 20.5. Above the plasmid is a segment of linear aardvark DNA that was synthesized using PCR. Diagram your cloning procedure, and show what would happen to these two molecules during each step. Use one color for the aardvark DNA and its bases and another color for those of the plasmid. Label each step and all 5’ and 3’ ends.

Ethical considerations aside, if DNA-based technologies became widely used, discuss how they might change the way evolution proceeds, as compared with the natural evolutionary mechanisms that have operated for the past 4 billion years.

You hope to study a gene that codes for a neurotransmitter protein produced in human brain cells. You know the amino acid sequence of the protein. Explain how you might (a) identify what genes are expressed in a specific type of brain cell, (b) identify (and isolate) the neurotransmitter gene, (c) produce multiple copies of the gene for study, and(d) produce large quantities of the neurotransmitter for evaluation as a potential medication.

In a short essay (100–150 words), discuss how the genetic basis of life plays a central role in biotechnology.

The water in the Yellowstone National Park hot springs shown here is around 160°F (70°C). Biologists assumed that no species of organisms couldlive in water above about 130°F (55°C), so they were surprised to find several species of bacteria there, now called thermophiles(“heat-lovers”). You’ve learned in this chapter how an enzyme from one species, Thermus aquaticus, made feasible one of the most important DNA-based techniques used in labs today.Identify the enzyme, and indicate the value of its being isolated from a thermophile. Suggest other reasons why enzymes from this bacterium (or other thermophiles) might also be valuable.