DNA Cloning: An Overview
DNA CLONING: AN OVERVIEW
“Instant Notes in Molecular Biology (Turner et al., 2005): 105-109
Nurul Layyin H
Questions and Answers
- 1.
A wide variety of natural replicons have the properties required to allow them to act as ___________
- A.
DNA ligase
- B.
DNA probe
- C.
Alkaline phosphatase
- D.
Selectable marker
- E.
Cloning vectors
Correct Answer
E. Cloning vectorsExplanation
A wide variety of natural replicons have the properties required to allow them to act as cloning vectors. This means that these replicons have the necessary characteristics to be used as tools in the process of cloning DNA. Cloning vectors are DNA molecules that can carry foreign DNA fragments and replicate them inside a host organism. These vectors are essential for the production of multiple copies of a specific DNA sequence, which is a fundamental step in molecular biology research and biotechnology applications.Rate this question:
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- 2.
Most of the routine manipulations involved in gene cloning use Escherichia coli as _______
- A.
Cloning vectors
- B.
Genomic libraries
- C.
CDNA libraries
- D.
DNA probe
- E.
The host organism
Correct Answer
E. The host organismExplanation
Most of the routine manipulations involved in gene cloning use Escherichia coli as the host organism. Escherichia coli is a commonly used bacterium in molecular biology research due to its ease of cultivation and well-understood genetics. It can efficiently take up foreign DNA and replicate it, allowing for the production of multiple copies of the inserted gene. Additionally, E. coli can be easily manipulated and has a fast generation time, making it a convenient choice for genetic engineering experiments.Rate this question:
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- 3.
A gene which allows host cells containing the vector to be selected from amongst those which do not, usually by conferring resistance to a toxin.
- A.
Selectable marker.
- B.
DNA probe
- C.
Recombinant DNA
- D.
Ti plasmid
- E.
Plasmid
Correct Answer
A. Selectable marker.Explanation
A selectable marker is a gene that allows host cells containing the vector to be selected from those that do not. This is typically done by conferring resistance to a toxin. In this case, the selectable marker gene would enable the identification and selection of host cells that have taken up the vector, while those without the vector would not possess the resistance and would not survive the toxin.Rate this question:
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- 4.
An enzyme that joins two double-stranded DNAs end to end.
- A.
DNA ligase
- B.
RNase A
- C.
Alkaline phosphatase
- D.
Reverse transcriptase
- E.
Restriction enzyme
Correct Answer
A. DNA ligaseExplanation
DNA ligase is the correct answer because it is an enzyme that has the ability to join two double-stranded DNAs end to end. It does this by catalyzing the formation of phosphodiester bonds between the adjacent nucleotides of the DNA strands. This process is crucial in DNA replication, repair, and recombination, as it allows for the sealing of nicks or gaps in the DNA backbone. DNA ligase is commonly used in molecular biology techniques such as cloning and PCR to join DNA fragments together.Rate this question:
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- 5.
A set of clones containing DNA fragments derived directly from a genome.
- A.
Cloning vectors
- B.
Genomic librariy
- C.
CDNA libraries
- D.
DNA probe
- E.
The host organism
Correct Answer
B. Genomic librariyExplanation
A genomic library is a collection of clones that contains DNA fragments derived directly from a genome. This means that the clones in the library represent the entire genetic material of an organism. The other options, such as cloning vectors, cDNA libraries, DNA probe, and the host organism, do not necessarily contain the complete set of DNA fragments from the genome. Therefore, the correct answer is genomic library.Rate this question:
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- 6.
DNA copies (cDNA) are synthesized from the mRNA by _________
- A.
Subcloning
- B.
Hybridization
- C.
Digestion
- D.
Reverse transcription
- E.
Ligation
Correct Answer
D. Reverse transcriptionExplanation
Reverse transcription is the correct answer because it is the process by which cDNA is synthesized from mRNA. Reverse transcription involves the use of an enzyme called reverse transcriptase, which catalyzes the synthesis of cDNA using the mRNA template. This process is important in molecular biology research as it allows scientists to study gene expression and analyze the sequence of specific genes.Rate this question:
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- 7.
The joining together of artificially separated nucleic acid molecules via hydrogen bonding between complementary bases.
- A.
Subcloning
- B.
Hybridization
- C.
Digestion
- D.
Reverse transcription
- E.
Ligation
Correct Answer
B. HybridizationExplanation
Hybridization is the correct answer because it refers to the process of joining together artificially separated nucleic acid molecules through hydrogen bonding between complementary bases. This process allows for the formation of hybrid molecules by combining DNA or RNA strands from different sources. Hybridization is commonly used in molecular biology techniques such as DNA microarrays, Southern blotting, and in situ hybridization to study gene expression, genetic variation, and DNA-protein interactions.Rate this question:
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- 8.
Enzymes used in DNA cloning that removes phosphate from 5′-ends of double- or single-stranded DNA or RNA .
- A.
DNA ligase
- B.
RNase A
- C.
Alkaline phosphatase
- D.
Reverse transcriptase
- E.
Restriction enzyme
Correct Answer
C. Alkaline phosphataseExplanation
Alkaline phosphatase is the correct answer because it is an enzyme used in DNA cloning that removes phosphate from the 5′-ends of double- or single-stranded DNA or RNA. This dephosphorylation step is important in preventing self-ligation of the vector during cloning, as it removes the 5′-phosphate groups that can allow the vector to ligate to itself instead of the insert DNA. By dephosphorylating the vector, alkaline phosphatase helps ensure that the insert DNA can be successfully ligated into the vector without unwanted self-ligation occurring.Rate this question:
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- 9.
A labelled molecule used in hybridisation procedures.
- A.
Cloning vector
- B.
Genomic library
- C.
CDNA library
- D.
DNA probe
- E.
Plasmid
Correct Answer
D. DNA probeExplanation
A DNA probe is a labelled molecule used in hybridization procedures. It is a short, single-stranded DNA sequence that is complementary to a specific target DNA sequence. The probe is labeled with a detectable marker, such as a fluorescent dye or radioactive isotope, which allows it to bind to the target sequence and be visualized. DNA probes are commonly used in molecular biology techniques, such as Southern blotting and in situ hybridization, to detect and identify specific DNA sequences of interest.Rate this question:
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- 10.
An endonuclease that cuts DNA at sites defined by its recognition sequence.
- A.
DNA ligase
- B.
RNase A
- C.
Alkaline phosphatase
- D.
Reverse transcriptase
- E.
Restriction enzyme
Correct Answer
E. Restriction enzymeExplanation
A restriction enzyme is an endonuclease that cuts DNA at specific recognition sequences. It recognizes a specific sequence of nucleotides and cuts the DNA at that site, creating fragments. This enzyme is commonly used in molecular biology techniques such as DNA cloning and genetic engineering to manipulate DNA. DNA ligase, RNase A, alkaline phosphatase, and reverse transcriptase are not restriction enzymes and do not have the same function of cutting DNA at specific recognition sequences.Rate this question:
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- 11.
An enzyme that hydrolyses (degrades) single-stranded DNA.
- A.
S1 nuclease
- B.
RNase A
- C.
Alkaline phosphatase
- D.
Reverse transcriptase
- E.
Terminal transferase
Correct Answer
A. S1 nucleaseExplanation
S1 nuclease is the correct answer because it is an enzyme that specifically hydrolyses single-stranded DNA. This enzyme is commonly used in molecular biology research to degrade single-stranded DNA, making it useful for various applications such as mapping DNA-protein interactions or removing unwanted single-stranded DNA from a sample. RNase A is an enzyme that degrades RNA, alkaline phosphatase removes phosphate groups from DNA or RNA molecules, reverse transcriptase synthesizes complementary DNA from RNA, and terminal transferase adds nucleotides to the 3' end of DNA or RNA molecules.Rate this question:
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- 12.
An enzyme that adds nucleotide residues to the 3' terminus of an oligo- or polynucleotide.
- A.
S1 nuclease
- B.
RNase A
- C.
DNA polymerase I
- D.
Reverse transcriptase
- E.
Terminal transferase
Correct Answer
E. Terminal transferaseExplanation
Terminal transferase is the correct answer because it is an enzyme that adds nucleotide residues to the 3' terminus of an oligo- or polynucleotide. S1 nuclease, RNase A, DNA polymerase I, and reverse transcriptase do not have this specific function.Rate this question:
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- 13.
Applications of DNA cloning , often collected together under the term genetic engineering, are the following:
- A.
Engineering animals and plants, and gene therapy
- B.
Engineering proteins to alter their properties
- C.
DNA sequencing
- D.
Identification of mutations
- E.
All of the above
Correct Answer
E. All of the aboveExplanation
The correct answer is "All of the above". DNA cloning has various applications in genetic engineering, including engineering animals and plants, gene therapy, engineering proteins to alter their properties, DNA sequencing, and identification of mutations.Rate this question:
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- 14.
A collection of clones prepared from the mRNA of a given cell or tissue type,re presenting the genetic information expressed by such cells
- A.
Cloning vectors
- B.
Genomic library
- C.
CDNA library
- D.
DNA probe
- E.
The host organism
Correct Answer
C. CDNA libraryExplanation
A cDNA library is a collection of clones prepared from the mRNA of a given cell or tissue type, representing the genetic information expressed by such cells. This means that the cDNA library contains complementary DNA (cDNA) sequences that have been synthesized from the mRNA molecules present in the cell or tissue. These cDNA sequences represent the genes that are actively being expressed in the cell or tissue, providing a snapshot of the genetic information being utilized by the organism at a specific time. Therefore, the correct answer is cDNA library.Rate this question:
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- 15.
A vector for cloning very large pieces of DNA in yeast.
- A.
BAC
- B.
YAC
- C.
Ti plasmid
- D.
Plasmid
- E.
Phage M13
Correct Answer
B. YACExplanation
YAC stands for yeast artificial chromosome, which is a vector used for cloning very large pieces of DNA in yeast. YACs are capable of accommodating large DNA fragments, making them suitable for cloning and studying large genes or entire genomes. This vector is widely used in molecular biology research to clone and manipulate large DNA sequences in yeast cells.Rate this question:
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- 16.
An RNA-dependent DNA polymerase found in retroviruses, used in vitro for the synthesis of cDNA.
- A.
S1 nuclease
- B.
RNase A
- C.
DNA polymerase I
- D.
Reverse transcriptase
- E.
Terminal transferase
Correct Answer
D. Reverse transcriptaseExplanation
Reverse transcriptase is an RNA-dependent DNA polymerase found in retroviruses. It is used in vitro for the synthesis of cDNA, which is complementary DNA synthesized from an RNA template. This enzyme is able to convert the single-stranded RNA molecule into a double-stranded DNA molecule by catalyzing the synthesis of DNA using the RNA template. Reverse transcriptase plays a crucial role in the replication of retroviruses and is also widely used in molecular biology research for techniques such as reverse transcription polymerase chain reaction (RT-PCR).Rate this question:
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- 17.
The analysis of the fragmentation of the DNA with restriction enzymes.
- A.
Subcloning
- B.
Hybridization
- C.
Digestion
- D.
Restriction mapping
- E.
Transformation
Correct Answer
D. Restriction mappingExplanation
Restriction mapping is the correct answer because it involves the analysis of DNA fragmentation using restriction enzymes. Restriction enzymes cut DNA at specific sequences, creating fragments of different lengths. By digesting DNA with these enzymes and analyzing the resulting fragments, scientists can determine the locations of restriction sites and create a map of the DNA molecule. This technique is widely used in molecular biology to study gene structure, identify mutations, and analyze DNA sequences. Subcloning, hybridization, digestion, and transformation are all related techniques but do not specifically involve the analysis of DNA fragmentation with restriction enzymes.Rate this question:
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- 18.
Technique for separating nucleic acid molecules on the basis of their movement through a gel matrix under the influence of an electric field.
- A.
Subcloning
- B.
Hybridization
- C.
Gel electrophoresis
- D.
Ligation
- E.
The polymerase chain reaction
Correct Answer
C. Gel electrophoresisExplanation
Gel electrophoresis is a technique used to separate nucleic acid molecules based on their movement through a gel matrix under the influence of an electric field. The gel matrix acts as a sieve, allowing smaller molecules to move faster and travel further than larger molecules. This separation is crucial in various molecular biology applications, such as DNA fingerprinting, genetic mapping, and studying gene expression. Gel electrophoresis is a fundamental tool in molecular biology research and is widely used to analyze and characterize nucleic acids.Rate this question:
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- 19.
Plasmid of Agrobacterium tumefaciens that can be used to integrate DNA into the genome.
- A.
SV40
- B.
YAC
- C.
Ti plasmid
- D.
Baculovirus
- E.
Phage M13
Correct Answer
C. Ti plasmidExplanation
The Ti plasmid of Agrobacterium tumefaciens is capable of integrating DNA into the genome. This plasmid is commonly used in genetic engineering to introduce foreign DNA into plants. The Ti plasmid contains the T-DNA region, which can be transferred to plant cells and integrated into their genome. This integration allows for the stable expression of the foreign DNA in the plant, making the Ti plasmid an important tool in plant biotechnology.Rate this question:
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- 20.
An enzyme derived from a thermostable bacterium (Thermus aquaticus). Operates at 72°C and is reasonably stable above 90°C. Used in PCR
- A.
S1 nuclease
- B.
Taq DNA polymerase
- C.
DNA polymerase I
- D.
Reverse transcriptase
- E.
Exonuclease III
Correct Answer
B. Taq DNA polymeraseExplanation
Taq DNA polymerase is the correct answer because it is derived from Thermus aquaticus, a thermostable bacterium. This enzyme is able to operate at high temperatures, specifically 72°C, which is necessary for the Polymerase Chain Reaction (PCR) process. Additionally, Taq DNA polymerase is reasonably stable above 90°C, which is important for the denaturation step in PCR. Therefore, Taq DNA polymerase is the enzyme of choice for PCR due to its ability to withstand high temperatures and perform DNA amplification efficiently.Rate this question:
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