LS 7A Week 8 Launchpad 3e
Assume you are trying to insert a gene into a plasmid. Someone gives you a preparation of genomic DNA that has been cut with restriction enzyme X. The gene you wish to insert has sites on both ends for cutting by restriction enzyme Y. You have a plasmid with a single site for Y but not for X. Your strategy should be to:
cut the DNA again with restriction enzyme Y and insert these fragments into the plasmid cut with restriction enzyme Y
One characteristic of restriction enzymes is that they cut:
double-stranded DNA at specific sites.
What is the name for the enzyme activity that DNA polymerase has when it removes an improperly paired base from the end of the growing strand?
exonuclease activity
If a restriction site of AatII is 5′-GACGTC-3′ then 3′-GACGTC-5′ is also an AatII restriction site.
false
Telomerase is fully active in _____ and _____ cells, but almost completely inactive in _____ cells.
germ; stem; somatic
In contrast to linear DNA replication, circular DNA replication typically:
has a single origin of replication.
What is the name of the enzyme that separates the two strands of DNA during replication?
helicase
A DNA molecule is cut with two different restriction enzymes known to cleave it only once each. After gel electrophoresis, three different DNA fragments are detected. This result means that the original DNA molecule was:
linear.
Restriction enzymes cleave double-stranded DNA at the sites that show a particular type of symmetry; these sequences read the same on both strands and are called palindromes. Which of these sequences is not a palindrome?
5'-CCTCAGG-3'
A certain restriction enzyme X cleaves double-stranded DNA at the sequence shown, where the slash indicates where each strand is cleaved. 5'-AA/ATTT-3' 3'-TTTA/AA-5' Note that the cleavage results in a two-base pair single-stranded region at the 5' end that allows the cleaved ends to undergo base pairing. Which of the restriction enzymes that cleave double-stranded DNA (indicated by the slash) would produce overhanging ends able to pair with those produced by enzyme X?
5'-GG/ATCC-3' 3'-CCTA/GG-5'
The enzyme responsible for joining Okazaki fragments together during DNA replication is:
DNA ligase.
Why are primers needed for DNA replication?
DNA polymerase can only add nucleotides to an existing chain.
The correct sequence of steps in the eukaryotic cell cycle is:
G1→ S phase → G2→ mitosis → cytokinesis.
Which lane contains the DNA fragments produced by digestion of the plasmid with enzymes X and Y and Z?
Lane H
Below on the left is a diagram of double-stranded DNA of a circular plasmid from a bacterial cell. The positions X, Y, and Z are sites where the plasmid DNA is cleaved by restriction enzymes X, Y, and Z, respectively. The numbers are the size in kilobase pairs (kb) of the DNA regions between the restriction sites. On the right is a diagram of an electrophoresis gel. Lane 1 is the ladder, comprised of DNA fragments that range in size from 1-12 kb showing the position of each band size after electrophoresis. The other lanes are DNA bands observed from digestion of the plasmid with one of more of the restriction enzymes. Which lane contains the DNA fragments produced by digestion of the plasmid with enzyme X only?
Lane K
Which lane contains the DNA fragments produced by digestion of the plasmid with enzymes Y and Z?
Lane L
Which lane contains the DNA fragments produced by digestion of the plasmid with enzymes X and Y?
Lane M
Which lane contains the DNA fragments produced by digestion of the plasmid with enzymes X and Z?
Lane Q
Imagine that a doctor is culturing two types of cells: one from a malignant melanoma and one from a normal skin sample. How would you expect these two cell populations to differ?
Malignant melanoma cells would have active telomerases that constantly replenish and lengthen telomeres.
Why are telomeres a necessary component of linear chromosomes?
They maintain the length of a chromosome because DNA is shortened every time it is replicated.
Which strand is bound to more RNA primers to the left of the dotted line?
Top strand
A Southern blot is a technique that relies on hybridization of:
a nucleic acid probe to a complementary DNA.
The transformation step in creating bacteria genetically engineered to produce human proteins involves:
bacteria taking up the recombinant DNA in the form of the vectors.
A graduate student wants to create a recombinant DNA molecule and introduce this molecule into bacteria. What is the proper order of steps that he should follow?
choose plasmid/donor DNA → cut with restriction enzymes → join fragments via DNA ligase → transform bacteria
What features of DNA make it possible to make recombinant DNA in the lab?
All of these choices are correct.
You would like to ligate a fragment you cleaved with the restriction enzyme BamHI into a plasmid you cleaved with a different enzyme. Which of these enzymes' overhangs will be compatible with BamHI-generated fragments? BamHI sequence and cleavage site: 5' G^GATCC 3' (^ indicates cut site)
BclI 5' T^GATCA 3'
Where is DNA polymerase found to the left of the dotted line?
Both strands
RNA primase lays down an RNA primer to start DNA replication. If a mutation occurs that blocks the function of RNA primase, which synthesis of which strand is affected, and why?
Both strands are affected because all DNA polymerase can only add new nucleotides to a 3′-OH present at the end of the RNA primer.
Which of the strands use a template for DNA replication?
Both strands use a template during DNA replication.
The diagram below represents the replication bubble that has formed at a bidirectional origin of replication. The dotted line shows where replication first began. Use this diagram to answer the questions below. Where does lagging strand synthesis occur to the right of the dotted line?
Bottom strand
Consider a cell in which one of the proteins involved in DNA replication is altered. This alteration results in an increased occurrence of single-stranded DNA breaks in the newly synthesized DNA. Which protein function is most likely nonfunctional in this situation?
The fragment joining function of ligase
To cells that are defective in primer removal, you add fluorescent ribonucleotides when the cells are undergoing DNA replication. In this case, you observe that one strand glows more than the other not only near the replication fork but also at intervals along its length. Which strand glows in this way and why?
The lagging strand glows in this way because its RNA primers are required for each Okazaki fragment and are closely spaced.
Suppose you add fluorescent ribonucleotides to a cell undergoing DNA replication so that the RNA primers used in DNA synthesis glow when viewed with a fluorescent microscope. You notice that, near each replication fork, one strand glows more than the other. Which strand glows more, and why?
The lagging strand glows more because its RNA primer is nearer the replication fork.
Which does not occur after an incorrect base is removed by DNA polymerase?
The template and daughter strands are separated by helicase activity.
Which of the statements best describes the way you would engineer bacterial cells to produce a human protein?
Use a specific restriction enzyme to cleave both the donor DNA and the vector DNA.
Which statement about DNA replication is true?
When DNA polymerase reaches a primer from prior DNA synthesis, the primer is removed and replaced with DNA.
In the diagrams below, the horizontal lines represent double-stranded DNA and the numbers are regions within the DNA. The sites labeled W, X, Y, and Z denote the locations of cleavage sites for the restriction enzymes W, X, Y, and Z, respectively. The DNA at the left is located in a bacterial plasmid vector and that on the right is located in chromosomal DNA. Each restriction enzyme produces ends that can pair with the ends of other DNA fragments produced by the same restriction enzyme but not with the ends produced by any of the other enzymes shown. A researcher plans to produce a clone with the genomic DNA inserted into the vector in the orientation 1-4-3-2. Which two enzymes should be used to cleave the vector and the chromosomal DNA to produce complementary ends that will lead to this result?
X and Y
A eukaryotic chromosome has _____ origin(s) of replication, and a bacterial chromosome has ______ origin(s) of replication.
many; one
The division of genetic material in a eukaryotic cell is called:
mitosis.
Which of the options is circled in this electron micrograph?
one double-stranded DNA molecule
Each DNA parent strand within a replication bubble acts as a template strand that produces:
one leading strand and one lagging strand.
As a piece of linear DNA is replicated, the leading strand will have _____ RNA primer(s) and the lagging strand will have _____ RNA primer(s).
one; many
What is the process of "proofreading" by DNA polymerase?
removing an improperly paired base and replacing it with the proper one
What is the name of the class of enzymes that recognizes and cuts a specific sequence of DNA?
restriction enzymes
Which of the choices is not needed for DNA replication?
ribosomes
If you were able to find a drug that could inhibit the reactivation of telomerase activity in cancer cells, the cancer cells would:
slowly erode their chromosome ends.
In DNA replication, each individual parent strand acts as a _____ strand for the synthesis of a _____ strand.
template; daughter
n which strand is DNA synthesis discontinuous, occurring in fragments that are later connected?
the lagging strand
On which strand are new nucleotides being added in the same direction as the replication fork is opening?
the leading strand
Refer to Animation: DNA Replication. What is the first thing to occur in DNA replication?
the strands of the DNA double helix are separated
What is the function of the enzyme DNA polymerase?
to synthesize a strand of DNA using DNA as a template
Restriction enzymes recognize certain DNA sequences and some of them will cut straight through, and others will leave an overhang at both ends of the cut.
true
A skin cell in G2 of interphase has _____ as much DNA as it had in G1.
twice
Refer to Animation: Proofreading in DNA Replication. How frequently is an improper base incorporated into the growing daughter strand?
very rarely
The parallel lines shown represent the paired strands of a DNA double helix. If this molecule undergoes one round of replication, which ends are shorter in the daughter molecules than in the parental molecules?
w and z
When we say that DNA replication is semiconservative, we mean that:
when DNA is replicated each new double helix contains one parental strand and one newly synthesized daughter strand.
