Ch 5 - Exploring Genes and Genomes
Imagine that the last 10 bases of a template end with 3′-ACCGGGTTTT-5′. Consider the sequencing products that will be made using these last 10 bases as a template; how many of these 10 sequencing products will contain dideoxy-T? 4 10 1 2 3
1 Beginning at the 5′ end the dideoxy sequence will be 3′-AAAACCCGGT-5′; there is one thymine at the end of the sequence; therefore, there is only one ddT.
If two 25-nucleotide primers are used for a 100-base pair DNA sequence in PCR, how long is the final amplified product? 48 base pairs 50 base pairs 100 base pairs 75 base pairs 25 base pairs
100 base pairs Primers do not add to the length of the DNA that is being amplified. PCR primers anneal to specific areas of DNA.
Which one of the following best describes a siRNA? approximately 90 bases of single-stranded RNA, folded into a cloverleaf shape 19 bp of double-stranded RNA with two unpaired bases at each 5′ end 6 bp of double-stranded palindromic RNA It is the same length as the corresponding gene's mRNA, but with the complementary sequence. It is the same length as the corresponding gene's 3′untranslated region, but with the complementary sequence.
19 bp of double-stranded RNA with two unpaired bases at each 5′ end A double-stranded RNA molecule is cleaved into 21-bp fragments by the enzyme Dicer to produce siRNAs. These siRNAs are incorporated into the RNA-induced silencing complex (RISC), where the single-stranded RNAs guide the cleavage of mRNAs that contain complementary sequences.
In a microarray experiment in which the expression of 10,000 genes was measured in five different tumor samples, how many data points would be plotted in a "heat map" representation of this data? 8000 10,000 5 2000 50,000
50,000 If there are 10,000 genes and 5 samples. There will be a total of 50,000 genes.
Which of the following sequences would MOST likely be cut by a restriction enzyme? 5′ TGCTGC 3′ 5′ AUGCAU 3′ 5′ GGCGCC 3′ 5′ CCGGCG 3′ 5′ GAGGTG 3′
5′ GGCGCC 3′ The staggered cuts made by this enzyme produce complementary single-stranded ends, which have specific affinity for each other and hence are known as cohesive or sticky ends.
Primers are normally approximately 25 bases in length, but for simplicity for this question assume that they are 5 bases in length. If one strand that you are trying to PCR-amplify has the sequence 5′-AGGTC-3′, which one of the following would be a suitable primer sequence? 5′-GACCU-3′ 5′-GACCT-3′ 5′-TCCAG-3′ 5′-AGGTC-3′ 5′-CTGGA-3′
5′-GACCT-3′ These base pairs match up, and the 5′ and 3′ match up well.
Which one of the following temperatures is best suited for the strand separation step in PCR? 54 C 37 C 95 C 72 C 110 C
95 C The first step in PCR is the denaturation step. For the strand to separate, a temperature of 95 C is optimal.
To identify a new gene, a DNA probe can be prepared from information about a short stretch of amino acids that are coded for by that gene. How many oligonucleotide probes would need to be synthesized to be sure that one complemented the DNA sequence that generated this stretch of amino acids? Arg - His - Met - Tyr - Phe - Trp - Phe 16384 96 65536 256 32768
96 The different possible arrangements of the seven amino acids led to 96 possible oligonucleotide probes to be synthesized to be sure of one complementary sequence. To search for Arg-His, there are 6 codons for Arg and 2 for His; thus 12 oligonucleotide probes would be required
If a DNA fragment lacks a desired restriction enzyme cut site, what can be done to introduce the desired sequence? Use a T4 phage. A linker can be covalently added. Add a 200-nucleic acid sequence to ensure the desired sequence is included. Nothing can be done. A new DNA fragment should be used Join fragments together using a helicase.
A linker can be covalently added. A linker adds the necessary bases so the restriction enzyme can cut at the appropriate site.
Why is a small amount of the 2′,3′-dideoxy analog of a nucleotide added to sequencing reactions? As a chain terminator, it lacks the 3′-hydroxyl needed to form the next phosphodiester bond. As a chain initiator, it can be incorporated by DNA polymerase without a primer. As a chain initiator, it contains the 5′-hydroxyl necessary at the beginning of a newly synthesized DNA strand. As a chain terminator, it lacks a base. As a chain terminator, it contains an ester group that blocks DNA polymerase from proceeding along the template strand.
As a chain terminator, it lacks the 3′-hydroxyl needed to form the next phosphodiester bond. It is a chain terminator because it lacks the hydroxyl needed to bond to continue the sequencing reaction; therefore, the amplification ceases.
Which one of the following is true regarding cosmids? A cosmid is the protein coat of a viral particle. Cosmids are found in nature when viruses are entering the lysogenic pathway. A cosmid is a hybrid formed from bacterial genomic DNA and a plasmid. Cosmids are suitable vectors for large inserts (e.g., 45 kilobases in length). Cosmids always contain the gene β-galactosidase.
Cosmids are suitable vectors for large inserts (e.g., 45 kilobases in length). Among the variety of lambda mutants that have been constructed for use as cloning vectors, one of them, called a cosmid, is essentially a hybrid of lambda phage and a plasmid that can serve as a vector for large DNA inserts.
Which of the following is the main difference between DNA and cDNA in eukaryotes? DNA contains exons and introns, whereas cDNA only contains introns. cDNA contains uracil, whereas DNA does not. DNA contains uracil, whereas cDNA does not. DNA contains introns, whereas cDNA does not. DNA contains exons, whereas cDNA does not.
DNA contains introns, whereas cDNA does not. DNA contains introns and cDNA does not contain introns. That is the difference between collection of DNA and cDNA in eukaryotes.
Next-generation sequencing methods tend to employ this enzyme: polynucleotide kinase sequenase DNA ligase a member of the restriction endonuclease family DNA polymerase
DNA polymerase Individual DNA fragments are amplified by PCR on a solid-support small region of a glass slide such that clusters of identical DNA fragments are distinguishable by high-resolution imaging. These fragments then serve as templates for DNA polymerase, where the addition of nucleotide triphosphates is converted to a signal that can be detected in a highly sensitive manner.
What type of enzyme is required for Sanger sequencing? Kinase A DNAse DNA polymerase DNA ligase a restriction enzyme
DNA polymerase Measurement of base incorporation in next-generation sequencing methods relies on the detection of the various products of the DNA polymerase reaction. Reversible terminator sequencing measures the nucleotide incorporation in a manner similar to Sanger sequencing.
How does DNA ligase function on DNA strands? DNA replication and DNA repair DNA ligase is only available in nature. Ligases can join any two 3′ DNA fragments with "blunt" ends. It connects Okinawa fragments. catalyzing the formation of a phosphoester bonds
DNA replication and DNA repair DNA ligase replicates and repairs DNA and can join any two complementary DNA fragments with cut ends. It can also catalyze the formation of a phosphodiester bond and connect Okazaki fragments.
Which technique would be used to determine the precise nucleotide composition of a segment of DNA? western blotting solid-phase synthesis of nucleic acids polymerase chain reaction (PCR) Southern blotting DNA sequencing
DNA sequencing This is the process of determining the order of nucleotides along chromosomes.
A complementary DNA (cDNA) library is a collection of _____ sequences representing all of the _____ expressed by a cell. DNA; DNA cDNA; RNA DNA; RNA cDNA; DNA DNA; mRNA
DNA; mRNA A cDNA library is a collection of DNA sequences that represents all of the mRNA in a cell.
When a double-stranded RNA molecule is introduced into an appropriate cell, the RNA is cleaved into fragments by an enzyme referred to as _____, thus producing small interfering RNA. RNA ligase RNA phosphatase RNase RNA restriction endonuclease Dicer
Dicer When a double-stranded RNA molecule is introduced into an appropriate cell, the RNA is cleaved by the enzyme Dicer into fragments approximately 21 nucleotides in length.
After staining the double-stranded DNA in an agarose gel with ethidium bromide, what do you need to do in order for the DNA to fluoresce bright orange? Expose it to ultraviolet light. Wait overnight. Expose it to orange light. Heat it. Keep it in darkness.
Expose it to ultraviolet light. Bands of radioactive DNA in gels can be visualized by autoradiography. A gel can be stained with a dye such as ethidium bromide, which fluoresces an intense orange under irradiation with ultraviolet light when bound to a double-helical DNA molecule. A band containing only 10 ng of DNA can be readily seen.
Why does gene disruption give clues to gene function? DNA heterogeneity maximization Foreign RNA inserted into a cell can disrupt any gene that is partly homologous. Regions of strong sequence similarity exchange segments of DNA through homologous recombination. Specific genes can be targeted if their nucleotide sequences are known. Foreign DNA inserted into a gene can disrupt the normal function of that gene and the resulting disruption of function can be observed.
Foreign DNA inserted into a gene can disrupt the normal function of that gene and the resulting disruption of function can be observed. When foreign DNA is inserted into the cell it disrupts the normal function of that gene, resulting in gene function that can be observed.
Which of the following is TRUE of the RNA interference mechanism? Dicer cleaves incoming single-stranded RNA into smaller fragments. The RNA-induced silencing complex (RISC) produces small interfering RNA (siRNA). Fully assembled RISC cleaves mRNA molecules that contain exact binding sites for RNase. Once RISC binds, it unwinds the strands of the siRNA and cleaves the guide strand. Fully assembled RISC cleaves mRNA molecules that contain exact complements of the guide strand.
Fully assembled RISC cleaves mRNA molecules that contain exact complements of the guide strand. The fully assembled RISC cleaves mRNA molecules that contain exact complements of the guide-strand sequence. Thus, levels of such mRNA molecules are dramatically reduced.
To study the physiological effects of a mutation in a human gene, which of the following would be the MOST appropriate? Microinject a plasmid containing the mutant gene into mammalian cells. Infect insect cells with a baculovirus containing the mutant gene. Generate a transgenic mouse carrying the mutated gene and observe the symptoms exhibited by the mouse. Infect human cells with a retrovirus containing the mutant gene. Isolate the mutant gene and express the gene in bacteria.
Generate a transgenic mouse carrying the mutated gene and observe the symptoms exhibited by the mouse. Mice harboring gene knockouts can be utilized as animal models for known human genetic diseases, enabling the assessment of new potential therapies for treatment.
Which of the following restriction enzymes cleave at the twofold axis of symmetry of their target DNA? BamHI EcoRI XhoI HaeIII HhaI
HaeIII HaeIII cleaves on the axis of symmetry, leaving no overlap or "sticky end."
Which of the following strategies might work to prevent the expression of a disease-associated gene? Introduce double-stranded RNA having homology to the disease gene that would be cleaved by DNA ligase, bound by RISC, and then used to recognize and bind to the mRNA of the disease-causing gene, preventing translation of the mRNA. Introduce single-stranded RNA having homology to the disease gene that would then be cleaved by Dicer, bound by RISC, and then used to recognize and cleave the mRNA of the disease-causing gene. Introduce double-stranded RNA having homology to the disease gene that would then be cleaved by Dicer, bound by RISC, and then used to recognize and bind to the mRNA of the disease-causing gene, preventing translation of the mRNA. Introduce a plasmid containing a highly expressed copy of the disease gene into the affected individual. Introduce double-stranded RNA having homology to the disease gene that would then be cleaved by Dicer, bound by RISC, and then used to recognize and cleave the mRNA of the disease-causing gene.
Introduce double-stranded RNA having homology to the disease gene that would then be cleaved by Dicer, bound by RISC, and then used to recognize and cleave the mRNA of the disease-causing gene. Homologous recombination is when two DNA molecules with strong sequence similarity exchange segments of DNA. They are flanked by sequences that have high homology to a particular region of genomic DNA and they will yield the transfer of the foreign DNA into the genome. When a double-stranded RNA molecule is introduced into an appropriate cell, the RNA is cleaved by the enzyme Dicer into fragments. This would have the desired effect of reducing or eliminating the expression.
What purpose is served by a polylinker region within a plasmid? It contains multiple reporter genes and multiple unique restriction sites. It contains multiple bacterial resistance genes and multiple unique restriction sites. It contains multiple unique restriction sites. It contains multiple reporter genes. It contains multiple bacterial resistance genes.
It contains multiple unique restriction sites. These polylinkers can be cleaved with a variety of restriction enzymes or combinations of enzymes, providing great versatility in the DNA fragments that can be inserted.
Which one of the following BEST indicates where the vector "bacteriophage lambda" originates? bacteria viruses archaea all eukaryotes (but no prokaryotes) yeast (but not other eukaryotes)
Lambda DNA comes from a virus called phage lambda. This virus is harmless to man.
_____ is synthesized by bacteria-harboring plasmids that contain DNA complementary to mRNA. Exons Bacteria Proinsulin Insulin Yeast artificial chromosome
Proinsulin cDNA has many applications beyond the generation of genetic libraries. The overproduction and purification of most eukaryotic proteins in prokaryotic cells necessitates the insertion of cDNA into plasmid vectors.
Following processing by Dicer, with which complex do siRNAs associate? pre-initiation complex (PIC) spliceosome complex (SPC) pre-interference complex (PIC) RNA-integrating suppression complex (RISC) RNA-induced silencing complex (RISC)
RNA-induced silencing complex (RISC) These siRNAs are incorporated into the RNA-induced silencing complex (RISC), where the single-stranded RNAs guide the cleavage of mRNAs that contain complementary sequences.
The FIRST eukaryotic genome to be completely sequenced was Haemophilus influenzae. Yersinia pestis. Saccharomyces cerevisiae. Drosophila melanogaster. Caenorhabditis elegans.
Saccharomyces cerevisiae. The first eukaryotic genome to be completely sequenced was that of baker's yeast, Saccharomyces cerevisiae, in 1996. This yeast genome comprises approximately 12 million base pairs, distributed on 16 chromosomes, and encodes more than 6000 proteins.
Cleavage of target DNA by the CRISPR-Cas system of which bacterium requires only a single protein, the nuclease Cas9, and an engineered, single-stranded guide RNA? Haemophilus influenzae Streptococcus pyogenes Caenorhabditis elegans Drosophila melanogaster Saccharomyces cerevisiae
Streptococcus pyogenes Streptococcus pyogenes is the source of the CRISPR-Cas9 system.
How can "insertional inactivation" be used to a researcher's advantage? It allows expression of an inhibitor of translation. Successful insertion of a gene of interest will prevent expression of a reporter gene. Successful insertion of a gene of interest will allow expression of a reporter gene. It allows expression of a toxin that disrupts cell division. It allows expression of a protein that cuts DNA at specific sites.
Successful insertion of a gene of interest will prevent expression of a reporter gene. For example, in the pUC18 vector, insertion of a DNA fragment into the polylinker region disrupts the lacZα gene.
What is the benefit of using complementary DNA (cDNA) reverse transcribed from mature mRNA when creating a DNA library from a eukaryotic organism? Only DNA that has been reverse transcribed can be introduced into a vector. In bacteria, only mRNA produced from cDNA will contain a 5′ cap and a 3′ polyadenylation signal. Introduction of cDNA will avoid inappropriate processing of the gene by prokaryotic-splicing machinery. The introns are already removed, and thus the gene product can be expressed in bacteria that lack the mRNA-capping machinery. The introns are already removed, and thus the gene product can be expressed in bacteria that lack the splicing machinery.
The introns are already removed, and thus the gene product can be expressed in bacteria that lack the splicing machinery. cDNA transcribed from mature mRNA has the introns already removed so when it is in bacteria that does not have the splicing enzymes, it produces the proteins from the genes on the mRNA.
Which one of the following is true regarding Southern blotting? The probe is RNA and it hybridizes to an RNA target. The probe is an antibody and it hybridizes to a protein target. The probe is RNA and it hybridizes to a DNA target. The probe is single-stranded DNA and it hybridizes to a DNA target. The probe is single-stranded DNA and it hybridizes to an RNA target.
The probe is single-stranded DNA and it hybridizes to a DNA target. It is a procedure for identifying sequences of DNA on a gel, then transferring it to a second medium to detect hybridization to be carried out.
The figure below shows the results of a quantitative PCR experiment on seven different cDNAs. Which of the following statements is TRUE? Two graphs are shown. The first graph shows the curves obtained in fluorescence monitoring in qPCR. The second graph shows the relationship between CT values and the starting quantity of DNA. The second graph has X axis labeled as Starting quantity ranging from ten to the power of 0 to 10 to the power of six, in increments of orders of magnitude and Y axis is labeled as CT ranging from 10 to 35, in increments of five. There are six points on the graph and a line with a negative slope is drawn connecting the points. The points at each end of the line have values of ten to the power of 0 on the X axis and 30 on the Y axis, and of ten to the power of five on the X axis and 15 on the Y axis, respectively. There are points in between these two points for each of the values on the X axis scale; they form an almost perfect line between the points at each end of the graph, with the CT value decreasing linearly as the starting quantity increases. The sample depicted in purple crossed the threshold at 22 cycles. The sample depicted in dark green had the greatest threshold. The sample depicted in red had the greatest number of copies of the original cDNA template. The sample depicted in blue had a CT larger than that of the sample depicted in dark green. The sample depicted in pink had more original copies of cDNA than the sample depicted in purple.
The sample depicted in red had the greatest number of copies of the original cDNA template. In qPCR, fluorescence is monitored in the course of PCR amplification to determine CT, the cycle at which this signal exceeds a defined threshold. Each color represents a different starting quantity of DNA. Red had the greatest number of cDNA copies.
When creating a DNA probe from a known protein sequence, which one of the following best indicates why peptide sequences containing methionine and tryptophan are preferred? Both of these amino acid side chains have an expected neutral charge at pH 7. Both of these amino acids are classified as hydrophobic. Both of these amino acids are encoded by the same codon. These amino acids are each encoded by six different codons. These amino acids are each encoded by one codon.
These amino acids are each encoded by one codon. When creating a DNA probe, a single peptide sequence can be encoded by a number of different oligonucleotides. Peptide sequences containing tryptophan and methionine are preferred, because these amino acids are specified by a single codon, whereas other amino acid residues have between two and six codons.
Each lane shows DNA that was cleaved with a different restriction enzyme. Different restriction enzymes are used to make DNA fragments, and those fragments travel through the gel to the positive end. The smaller fragments move more quickly toward the positive end.
This figure shows the result of a restriction digest on the DNA from the virus SV40. Which of the following statements is TRUE? The photograph is rectangular with a black background, with different points on its horizontal axis numbered from one to eight, representing the lanes on the DNA gel. For each of these lanes, there is a vertical column on the gel with a series of small, horizontal, white bands of DNA across it. There is a scale on the vertical axis of the photograph, showing the unit bp. Near the top of the scale, the value 5000 is indicated, close to the middle of the scale, the number 1000 is indicated, and closer to the bottom of the scale, the number 500 is indicated. There is an un-numbered lane with DNA bands of known sizes at the very left of the photograph. The DNA in each lane was cleaved the same number of times. Each lane shows DNA that was cleaved with a different restriction enzyme. The same restriction enzyme was used to generate the samples in lanes A, B, and C, but the gel for lane C was allowed to run for a longer period of time. The same restriction enzyme was used to generate the samples in lanes A, B, and C; they were just incubated for longer periods of time. Lane A shows the DNA without any restriction enzyme added, whereas lanes B and C show the result of incubation with two different restriction enzymes.
For what reason would you most likely choose to perform electrophoresis on DNA fragments using a polyacrylamide gel instead of an agarose gel? Your DNA fragments are quite short. Your sample contains DNA wrapped around proteins. Your DNA sample is derived from a prokaryote. Your DNA sample has been stained with a fluorescent dye. You believe that your DNA sample is contaminated with RNA.
Your DNA fragments are quite short. DNA is negatively charged, so the different sizes of the fragments will move faster or slower through the gel to the positively charged side of the gel to yield information.
Which one of the following BEST gives an example of a "reporter gene"? a gene that encodes multiple proteins that are involved in the same biochemical pathway a gene that encodes an enzyme that requires ATP a gene that encodes an isozyme a pseudogene a gene that encodes a fluorescent protein
a gene that encodes a fluorescent protein A reporter gene is a gene that is attached to a sequence of another gene of interest. When the gene of interest is expressed, the fluorescent protein is also expressed, acting as a 'reporter' of the expression for the scientist.
Which one of the following best describes a restriction-fragment-length polymorphism (RFLP)? a mutation that prevents an individual's DNA from being copied during cell division an individual who produces mutated forms of restriction enzymes a mutation that changes a genomic sequence so that It can no longer be recognized and cut by a particular restriction enzyme an individual with fragmentation in his or her chromosomal DNA an individual who is restricted in his or her ability to produce restriction enzymes at high levels
a mutation that changes a genomic sequence so that It can no longer be recognized and cut by a particular restriction enzyme RFLPs are polymorphisms within restriction sites that change the sizes of DNA fragments produced by the appropriate restriction enzyme.
Which feature of an expression vector is particularly helpful if one wishes to affinity-purify the protein product expressed from the insert ligated into that vector? a gene for antibiotic resistance a polylinker region containing sequences for multiple restriction enzymes an origin of replication a tag sequence flanking the insertion site a reporter gene
a tag sequence flanking the insertion site An expression vector is a type of cloning vector. An expression vector has a promoter sequence to drive transcription. Through affinity-purifying, the protein of interest is purified by its specific binding properties to an immobilized ligand.
Which one of the following is the best choice of vector for an extremely large insert (e.g., one megabase)? a lambda phage a plasmid a bacterial artificial chromosome a cosmid a yeast artificial chromosome
a yeast artificial chromosome YACs contain a centromere, an autonomously replicating sequence. Inserts as large as 1000 kb can be cloned into YAC vectors.
DNA ligase catalyzes a specific reaction. What does it require for the reaction? inorganic phosphate adenosine diphosphate a restriction enzyme a plasmid with an additional enzyme adenosine triphosphate
adenosine triphosphate ATP are consumed by DNA ligase to join the phosphate backbone of DNA with blunt or compatible cohesive ends.
A vector can be useful in recombinant DNA methods if it
allows rapid insertion of DNA fragments of interest. The DNA fragment is isolated; then it is cleaved with a restriction enzyme. The vector DNA is gathered and placed together in the cell and allowed to replicate to produce a protein.
Which one of the following best describes where restriction enzymes are expressed in nature? vertebrates yeast plants They are not expressed in nature. bacteria
bacteria These enzymes are found in bacteria and archaea and provide a defense mechanism against invading viruses.
Which one of the following methods is used to express proteins in insect larvae? retrovirus baculovirus calcium phosphate precipitation microinjection vaccinia virus
baculovirus Baculovirus infects insects cells, which can be conveniently cultured. Insect larvae infected with this virus can serve as efficient protein factors
The guide strand of the siRNA must _____ to its mRNA target. induce mutations repel be complementary be random when compared be identical
be complementary These siRNAs are incorporated into the RNA-induced silencing complex (RISC), where the single-stranded RNAs guide the cleavage of mRNAs that contain complementary sequences.
What is the purpose of the dimethoxytrityl (DMT) group during the synthesis of DNA? blocks the 3′ oxygen links the base to the resin blocks the 5′ oxygen activates the 5′ oxygen to facilitate coupling activates the 3′ oxygen to facilitate coupling
blocks the 5′ oxygen Dimethoxytrityl (DMT) is added to the growing chain of deoxyribonucleoside 3′-phosphoramidite to protect the 5′-oxygen atom from any bonding.
Which one of the following best describes the role of DNA ligase? breaking phosphodiester bonds breaking glycosidic bonds between deoxyribose and bases in DNA copying DNA sequence adding missing bases to double-stranded DNA catalyzing phosphodiester bond formation
catalyzing phosphodiester bond formation DNA ligase catalyzes the forming of bonds between two complementary strands to form DNA.
Disruption of the regulatory gene myogenin generates normal animals if one copy of the gene has been knocked out. generates normal animals if both copies of the gene have been knocked out. causes animals to die at birth if both copies of the gene are knocked out. it has no effect on the animal. causes animals to die at birth if one copy of the gene has been knocked out.
causes animals to die at birth if both copies of the gene are knocked out. When both copies of the gene for the regulatory protein myogenin are disrupted, an animal dies at birth because it lacks functional skeletal muscle.
What advantage do we gain comparing an organism like puffer fish with the human genome? understanding the phenotype of the different puffer fish Puffer fish have the intergenic DNA in abundance. comparison of genes previously unrecognized human genes over 1 gigabase pairs of sequences to compare to human genes The puffer fish genome is larger than the human genome.
comparison of genes previously unrecognized human genes The puffer fish genomes include fewer than 400 megabase pairs, one-eighth of the number in the human genome, yet the puffer fish and human genomes contain essentially the same number of genes. Comparison of the genomes of these species with that of humans revealed more than 1000 formerly unrecognized human genes.
Which of the following substrates does the Dicer enzyme prefer? single-stranded RNA single-stranded DNA RNA-DNA hybrid double-stranded DNA double-stranded RNA
double-stranded RNA A double-stranded RNA molecule is cleaved into 21-bp fragments by the enzyme Dicer to produce siRNAs. These siRNAs are incorporated into the RNA-induced silencing complex (RISC), where the single-stranded RNAs guide the cleavage of mRNAs that contain complementary sequences.
A mixture of DNA fragments can be separated by _____ on an agarose gel. The DNA will run toward the _____ electrode with the larger DNA fragments migrating more _____ than the smaller fragments. sequencing; positive; quickly electrophoresis; positive; slowly electrophoresis; negative; slowly sequencing; negative; slowly electrophoresis; positive; quickly
electrophoresis; positive; slowly DNA fragments are separated using electrophoresis, during which DNA fragments will migrate toward the positive electrode. Larger fragments interact more with the agarose, travelling more slowly through the gel than smaller fragments.
Which of these components, used in agarose gel experiments, fluoresce intense orange when bound to a double-helical DNA molecule? bromophenol blue methyl orange ethidium bromide xylene cyanol coomassie blue
ethidium bromide The dye is bound to the end of the DNA and replicated with the DNA. The benzene rings on the dye are excited by energy from lasers and release visible photons. When ethidium bromide is excited it emits bright orange.
Which of the following describes a step necessary for the successful production of recombinant DNA? generation of cohesive 3′ends on both the DNA fragment of interest and binding them together for a cohesive DNA strand for replication cleavage of two DNA fragments and covalently binding them together with a phosphate bond cleavage of the DNA fragment of interest and the vector DNA with a different restriction enzyme cleavage of the DNA fragment of interest and the vector DNA with partner restriction enzymes generation of complementary cohesive ends on both the DNA fragment of interest and the vector DNA
generation of complementary cohesive ends on both the DNA fragment of interest and the vector DNA First, the DNA fragment is isolated; then it is cleaved with a restriction enzyme. The vector DNA is gathered and placed together in the bacterial cell and allowed to replicate to produce a protein.
Completion of the human genome sequencing project indicated that we have more than expected of each of the following EXCEPT: alternatively spliced transcripts genes short interspersed elements (SINEs) pseudogenes long interspersed elements (LINEs)
genes There are approximately 19,000-20,000 human genes. This is far lower than the initial predictions of 100,000.
Where are plasmids most commonly found? in large single-stranded RNA in mammals in bacteria in human genomes in mammalian mitochondria in fertilizer for use in plants
in bacteria They are mainly found in bacteria but also naturally in archaea and eukaryotes like yeast and plants.
In preparing cDNA of a transcribed gene, one needs a(n) _____ template. reverse transcriptase tRNA mRNA DNA polymerase rRNA
mRNA An mRNA template is used in synthesizing cDNA duplexes.
While the expression level of a gene can change in a cell over time, what will always be proportional to the protein level? triglycerides introns mRNA ribosomes tRNA
mRNA Messenger RNA that conveys genetic information from DNA to the ribosome, where it specifies the amino acid sequence of the gene expression. The mRNA requires proteins and codes the sequences so it will be proportional to the protein level. mRNA stability and rate of protein breakdown can both be affected by cellular needs.
Which technique can be used to generate a mutant protein that contains a single amino acid substitution? oligonucleotide-directed mutagenesis cassette mutagenesis DNA ligation polymerase chain reaction restriction enzyme digest
oligonucleotide-directed mutagenesis If we have a plasmid containing the gene or cDNA for the protein and we know the base sequence around the site to be altered, we can then substitute the base of our choice.
DNA polymerization alters which of the following? DNA polymerase primer hybridization to flanking sequences of the target pH mRNA structure cellulose wall structure
pH DNA polymerase drastically increases pH from 8.0 to approximately 9.0.
What modification of DNA fragments and decameric linkers can facilitate their joining by T4 ligase? methylation by methyltransferase phosphorylation by polynucleotide kinase phosphorylation by polyphosphatase methylation by mononucleotide kinase
phosphorylation by polynucleotide kinase First, the linker is covalently joined to the ends of a DNA fragment. For example, the 5′ ends of a decameric linker and a DNA molecule are phosphorylated by polynucleotide kinase and then joined by the ligase from T4 phage.
Which one of the following is capable of adding a 5′ phosphate to a DNA fragment if needed? polynucleotide kinase all restriction enzymes that produce blunt ends (but not those that produce sticky ends) all restriction enzymes DNA ligase all restriction enzymes that produce sticky ends (but not those that produce blunt ends)
polynucleotide kinase Polynucleotide kinase is bacteriophage enzyme that catalyzes the transfer of a phosphate from ATP to the free hydroxyl of the 5′ DNA.
Transgenic mice that carry the gene for a mutated form of SOD1 will NOT display which of the following symptoms associated with ALS? paralysis poor blood clotting rapid progression to death progressive weakness of voluntary muscles motor-neuron loss
poor blood clotting Mice with this mutation had degeneration in white matter sections of the spinal cord and loss of anterior horn neurons. Aggregates were seen in the anterior, posterior, and white matter continues brain loss and muscle degeneration. Due to muscle degeneration and atrophy, they will not live long enough to display poor blood clotting from human ALD gene SOD1.
What is the second step in a PCR cycle? DNA is synthesized primers anneal to 3′ ends of both target strands DNA ligase incorporation DNA bond breaking primers anneal to each other
primers anneal to 3′ ends of both target strands The annealing of primers is used to copy DNA. PCR uses two primers, each complementary to opposite strands of the DNA; the forward primer is complementary with the top strand and the reverse primer is complementary with the lowest strand. They were denatured by heating, then the primers are added at about 54 C to bind to each strand. One primer hybridizes to the 3′ end of the target on one strand, and the other primer hybridizes to the 3′ end on the complementary target strand.
What is present in an expression vector that differentiates it from a cloning vector? The only difference between an expression vector and a cloning vector is the overall size. a reporter gene promoter sequences to drive transcription a polylinker region antibiotic resistance genes
promoter sequences to drive transcription An expression vector is a type of cloning vector. The difference between them is that an expression vector has a promoter sequence to drive transcription.
The enzyme reverse transcriptase uses _____ as the template. single-stranded rRNA double-stranded DNA single-stranded mRNA double-stranded mRNA single-stranded DNA
single-stranded mRNA A reverse transcriptase used to generate complementary DNA from a single-stranded mRNA.
Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) can both be used for next-generation genome sequencing. injecting DNA into germ cells. generating cDNA from mRNA. site-specific genome editing. generating siRNAs.
site-specific genome editing. Using different specific binding domains, both zinc-finger nucleases and transcription activator-like effector nucleases can introduce double-stranded breaks very specifically into a gene of interest.
For screening a genomic library, which one of the following reagents or conditions can be used to both lyse the bacteria and create single-stranded DNA suitable for hybridization to the desired probe? sodium hydroxide lambda phage dilute sodium chloride ATP freezing temperature
sodium hydroxide Sodium hydroxide is used as an alkaline lysis buffer, which dissolves the cell membrane so that the inner components of the cell come out. Additionally, the most commonly used method for creating single-stranded DNA is alkaline denaturation, most commonly used in PCR, in which sodium hydroxide is used.
During the automated synthesis of a DNA strand by the phosphite triester p method, dichloroacetic acid is added at one step in order to remove: the 3′ phosphorus atom the DMT group, generating a free ester the β-cyanoethyl (βCE) group, generating a free hydroxyl the β-cyanoethyl (βCE) group, generating a free ester the DMT group, generating a free hydroxyl
the DMT group, generating a free hydroxyl The hydroxyl group is freed up by the addition of dichloroacetic acid, which leaves other protecting groups intact.
In fluorescence-based Sanger sequencing, which one of the following best indicates what specifically is being fluorescently tagged? the primer the dideoxynucleotides [Fluorescently-tagged dideoxynucleotides are similar to regular (deoxy) nucleotides; however, they lack a hydroxyl group on the 3' carbon of the sugar ring. The lack of that hydroxyl group prevents the addition of more nucleotides to the chain. ]] the deoxynucleotides the enzyme the template
the dideoxynucleotides [Fluorescently-tagged dideoxynucleotides are similar to regular (deoxy) nucleotides; however, they lack a hydroxyl group on the 3' carbon of the sugar ring. The lack of that hydroxyl group prevents the addition of more nucleotides to the chain. ]]
All of the following are TRUE statements regarding DNA that is cut by a restriction enzyme that produces staggered cuts EXCEPT the ends can be referred to as "blunt ends." there is more than one restriction enzyme that is capable of making staggered cuts. fragments with complementary staggered ends can be joined together using DNA ligase. the ends can be referred to as "sticky ends." the ends can be referred to as "cohesive ends."
the ends can be referred to as "blunt ends." The type of cuts these restriction enzymes make give them a different name.
Which part of the human genome was the first to be sequenced? the Y chromosome chromosome 22 chromosome 1 the mitochondrial genome the X chromosome
the mitochondrial genome Sanger and his coworkers determined the complete sequence of the 5386 bases in the genome of the φX174 DNA virus in 1977. This tour de force was followed several years later by the determination of the sequence of human mitochondrial DNA.
In electroporation, what is the purpose of applying electric pulses? introduction of double-stranded breaks in the in the genome introduction of base changes in the foreign DNA introduction of pores in the nucleus transiently increasing membrane permeability disruption of the integrity of the cell wall
transiently increasing membrane permeability Electric pulses are applied to a suspension of protoplasts and plasmid DNA. Because high-electric fields make membranes transiently permeable to large molecules, plasmid DNA molecules enter the cells. The cell wall is then allowed to reform, and the plant cells are again viable.
Plasmids and bacteriophage lambda are commonly used _____ for cloning in E. coli. translators transmutators vectors transfectors transformers
vectors A vector is a small piece of DNA that is taken from an organism. This DNA fragment can be inserted into another host for cloning.
A molecular oncologist is trying to determine if human papilloma virus has infected the cervical cancer cell line she is studying. She has isolated DNA from the cancer cells to test whether she can detect the viral genome within. Which of the following techniques would NOT be helpful in determining whether the isolated DNA contains a portion of the viral genome? Southern blotting solid-phase synthesis of nucleic acids western blotting gel electrophoresis PCR amplification
western blotting Western blotting is used to identify specific amino acid sequences in proteins.
The pUC18 expression vector contains a lacZ gene. Within the lacZ gene is a restriction site for EcoRI. To insert a DNA fragment into this vector a digestion of both vector and insert with EcoRI is followed by a ligation reaction. The product of this ligation is then introduced into bacteria. The bacteria are subjected to a β-galactosidase assay. What color will the bacterium be that uptake the vector with insertion? blue because the lacZ gene would be disrupted by the insertion A 50:50 mixture of white and blue bacteria would result. white because the lacZ gene would be disrupted by the insertion blue because the lacZ gene would not be disrupted by the insertion white because the lacZ gene would not be disrupted by the insertion
white because the lacZ gene would be disrupted by the insertion β-Galactosidase cleaves the synthetic substrate X-gal, releasing a blue dye. Therefore, disruption of the lacZ gene will result in white bacteria.
In a two-color microarray experiment using red and green fluorescent nucleotides, what is the expected spot color when a gene is highly expressed in both samples? green violet yellow red black
yellow mRNA is isolated from two samples. From these transcripts, cDNA is prepared in the presence of a fluorescent nucleotide, with a red label and a green label. The cDNA strands are separated, hybridized to the microarray, and the unbound DNA is washed away. Spots that are red indicate genes that are expressed more highly, while the green spots indicate reduced expression. Spots that are yellow indicate comparable expression at high levels in both samples.
