Complete Solutions Manual A First
Course in Differential Equations with
Modeling Applications Ninth
2. What feature is commonly seen in the sequences recognized by type II restriction
enzymes? - ANSWERS--Solution:
The recognition sequences are palindromic, and 4−8 base pairs long.
3. What normal role do restriction enzymes play in bacteria? How do bacteria protect
their own DNA from the action of restriction enzymes? - ANSWERS--Solution:
Restriction enzymes cut foreign DNA, such as viral DNA, into fragments. Bacteria
protect their own DNA by modifying bases, usually by methylation, at the recognition
sites.
9. Briefly explain how an antibiotic-resistance gene and the lacZ gene can be used to
determine which cells contain a particular plasmid. - ANSWERS--Solution:
Many plasmids designed as cloning vectors carry a gene for antibiotic resistance and
the lacZ gene. The lacZ gene on the plasmid has been engineered to contain multiple
unique restriction sites. Foreign DNAs are inserted into one of the unique restriction
sites in the lacZ gene of plasmids and the plasmids are transformed into E. coli cells
lacking a functional lacZ gene. Transformed cells are plated on a medium containing the
appropriate antibiotic to select for cells that carry the plasmid. The medium also
contains an inducer for the lac operon, so the cells express the lacZ gene, and X-gal, a
substrate for beta-galactosidase that will turn blue when cleaved by β-galactosidase.
The colonies that carry plasmid without foreign DNA inserts will have intact lacZ genes,
make functional β-galactosidase, cleave X-gal, and turn blue. Colonies that carry
plasmid with foreign DNA inserts will not make functional β-galactosidase because the
lacZ gene is disrupted by the foreign DNA insert. They will remain white. Thus, cells
, carrying plasmids with inserts will form white colonies. This is known as blue/white
selection.
4. Explain how gel electrophoresis is used to separate DNA fragments of different
lengths. - ANSWERS--Solution:
Gel electrophoresis uses an electric field to drive DNA molecules through a gel that acts
as a molecular sieve. The gel is an aqueous matrix of agarose or polyacrylamide. DNA
molecules are loaded into a slot or well at one end of the gel. When an electric field is
applied, the negatively charged DNA molecules migrate toward the positive electrode.
Shorter DNA molecules are less hindered by the agarose or polyacrylamide matrix and
migrate faster than longer DNA molecules, which must wind their way around obstacles
and through the pores in the gel matrix.
5. After DNA fragments have been separated by gel electrophoresis, how can they be
visualized? - ANSWERS--Solution:
DNA molecules can be visualized by staining with a fluorescent dye, such as ethidium
bromide, that intercalates between the stacked bases of the DNA double helix, and the
dye-DNA complex fluoresces when irradiated with an ultraviolet light source.
Alternatively, they can be visualized by attaching radioactive or chemical labels to the
DNA before it is placed in the gel.
6. What is the purpose of Southern blotting? How is it carried out? - ANSWERS--
Solution:
Southern blotting is used to detect and visualize specific DNA fragments that have a
sequence complementary to a labeled DNA probe. DNA is first cleaved into fragments
with restriction endonucleases. The fragments are separated by size via gel
electrophoresis. These fragments are then denatured and transferred by blotting onto
the surface of a membrane filter. The membrane filter now has single-stranded DNA
fragments bound to its surface, separated by size as in the gel. The filter is then
incubated with a solution containing a denatured, labeled probe DNA. The probe DNA
hybridizes to its complementary DNA on the filter. After washing away excess unbound
probes, the labeled probe hybridized to the DNA on the filter can be detected using the
appropriate methods to visualize the label. For radioactively labeled probes, the bound
probe is detected by exposure to X-ray film. Other probe labeling methods detect bound
probe using enzymatic reactions that generate luminescence or color.
7. Give three important characteristics of cloning vectors. - ANSWERS--Solution:
Cloning vectors should have:
(1) An origin of DNA replication so they can be maintained in a cell
(2) A gene, such as antibiotic resistance, to select for cells that carry the vector
(3) A unique restriction site or series of sites to where a foreign DNA molecule may
be inserted
10. Briefly explain how the polymerase chain reaction is used to amplify a specific DNA
sequence. - ANSWERS--First, the double-stranded template DNA is denatured by high
temperature. Then, primers corresponding to the ends of the DNA sequence to be
Course in Differential Equations with
Modeling Applications Ninth
2. What feature is commonly seen in the sequences recognized by type II restriction
enzymes? - ANSWERS--Solution:
The recognition sequences are palindromic, and 4−8 base pairs long.
3. What normal role do restriction enzymes play in bacteria? How do bacteria protect
their own DNA from the action of restriction enzymes? - ANSWERS--Solution:
Restriction enzymes cut foreign DNA, such as viral DNA, into fragments. Bacteria
protect their own DNA by modifying bases, usually by methylation, at the recognition
sites.
9. Briefly explain how an antibiotic-resistance gene and the lacZ gene can be used to
determine which cells contain a particular plasmid. - ANSWERS--Solution:
Many plasmids designed as cloning vectors carry a gene for antibiotic resistance and
the lacZ gene. The lacZ gene on the plasmid has been engineered to contain multiple
unique restriction sites. Foreign DNAs are inserted into one of the unique restriction
sites in the lacZ gene of plasmids and the plasmids are transformed into E. coli cells
lacking a functional lacZ gene. Transformed cells are plated on a medium containing the
appropriate antibiotic to select for cells that carry the plasmid. The medium also
contains an inducer for the lac operon, so the cells express the lacZ gene, and X-gal, a
substrate for beta-galactosidase that will turn blue when cleaved by β-galactosidase.
The colonies that carry plasmid without foreign DNA inserts will have intact lacZ genes,
make functional β-galactosidase, cleave X-gal, and turn blue. Colonies that carry
plasmid with foreign DNA inserts will not make functional β-galactosidase because the
lacZ gene is disrupted by the foreign DNA insert. They will remain white. Thus, cells
, carrying plasmids with inserts will form white colonies. This is known as blue/white
selection.
4. Explain how gel electrophoresis is used to separate DNA fragments of different
lengths. - ANSWERS--Solution:
Gel electrophoresis uses an electric field to drive DNA molecules through a gel that acts
as a molecular sieve. The gel is an aqueous matrix of agarose or polyacrylamide. DNA
molecules are loaded into a slot or well at one end of the gel. When an electric field is
applied, the negatively charged DNA molecules migrate toward the positive electrode.
Shorter DNA molecules are less hindered by the agarose or polyacrylamide matrix and
migrate faster than longer DNA molecules, which must wind their way around obstacles
and through the pores in the gel matrix.
5. After DNA fragments have been separated by gel electrophoresis, how can they be
visualized? - ANSWERS--Solution:
DNA molecules can be visualized by staining with a fluorescent dye, such as ethidium
bromide, that intercalates between the stacked bases of the DNA double helix, and the
dye-DNA complex fluoresces when irradiated with an ultraviolet light source.
Alternatively, they can be visualized by attaching radioactive or chemical labels to the
DNA before it is placed in the gel.
6. What is the purpose of Southern blotting? How is it carried out? - ANSWERS--
Solution:
Southern blotting is used to detect and visualize specific DNA fragments that have a
sequence complementary to a labeled DNA probe. DNA is first cleaved into fragments
with restriction endonucleases. The fragments are separated by size via gel
electrophoresis. These fragments are then denatured and transferred by blotting onto
the surface of a membrane filter. The membrane filter now has single-stranded DNA
fragments bound to its surface, separated by size as in the gel. The filter is then
incubated with a solution containing a denatured, labeled probe DNA. The probe DNA
hybridizes to its complementary DNA on the filter. After washing away excess unbound
probes, the labeled probe hybridized to the DNA on the filter can be detected using the
appropriate methods to visualize the label. For radioactively labeled probes, the bound
probe is detected by exposure to X-ray film. Other probe labeling methods detect bound
probe using enzymatic reactions that generate luminescence or color.
7. Give three important characteristics of cloning vectors. - ANSWERS--Solution:
Cloning vectors should have:
(1) An origin of DNA replication so they can be maintained in a cell
(2) A gene, such as antibiotic resistance, to select for cells that carry the vector
(3) A unique restriction site or series of sites to where a foreign DNA molecule may
be inserted
10. Briefly explain how the polymerase chain reaction is used to amplify a specific DNA
sequence. - ANSWERS--First, the double-stranded template DNA is denatured by high
temperature. Then, primers corresponding to the ends of the DNA sequence to be
