102 Questions
What is the main purpose of DNA cloning?
Generating recombinant DNA
What is the role of a vector in DNA cloning?
Serves as a carrier for the gene of interest
How do restriction endonucleases function in DNA cloning?
Recognize and cut DNA at specific sequences
What defines palindromic sequences in the context of DNA cloning?
Sequences that read the same forwards and backwards
How are blunt-ended fragments generated by restriction endonucleases?
By cutting at the same position on both strands
What distinguishes staggered (off-center) cuts by restriction endonucleases?
They cut the two DNA strands at different positions
What is the purpose of a DNA ligase in the process of making recombinant DNA?
To 'close' the plasmid
What characteristic of an expression vector distinguishes it from a regular vector?
It contains promoter and termination sequences
Why is it challenging to express proteins in bacteria?
They have internal disulfide bonds
What solution is proposed to overcome the challenges of protein expression in bacteria?
Employing eukaryotic systems like yeast
How can proteins be 'tagged' during the process of creation of protein hybrids?
Inserting a sequence of amino acids called tags
Which enzyme is crucial for sealing the DNA fragments after they hybridize in the process of making recombinant DNA?
DNA ligase
Which protein tag is detected using an antibody specific to a peptide sequence?
FLAG
What is a protein domain based on the provided text?
A region of the protein's polypeptide chain with a defined three-dimensional structure
Why is Green Fluorescent Protein (GFP) used according to the text?
For protein detection purposes
For which protein tag is purification achieved by binding to nickel or imidazole?
Poly-His
Which method involves separating proteins based on size in gel wells?
Gel electrophoresis
Which protein tag is used for protein purification purposes?
GST-tag
Which protein tag is associated with a ~220 amino acid protein and can be detected by fluorescence?
GFP
What makes a protein domain distinct according to the text?
It has a defined three-dimensional structure and folds independently
What is the purpose of the GST-tag in the context provided?
For purification of GST-tagged proteins
Which amino acid sequence is specific to the V5 protein tag?
GKPIPNPLLGLDST
What is the purpose of adding six histidines to a protein?
Purification using nickel or imidazole
What is the role of DNA ligase in the context of cloning?
Joins DNA ends by forming phosphodiester bonds
What is the purpose of cloning in genetics?
To make several copies of one thing
What is recombinant DNA technology?
Manipulating DNA for various purposes
Why are bacterial plasmids considered excellent vectors for cloning?
They replicate independently of the main bacterial genome
Which feature must plasmid cloning vectors have?
Presence of an antibiotic resistance gene
What is the main purpose of using restriction endonucleases in DNA cloning?
Generate sticky or blunt ends on DNA fragments
Which of the following accurately describes a palindromic sequence in the context of DNA cloning?
A sequence that reads the same from left to right as it does from right to left
In DNA cloning, what is the significance of generating sticky or cohesive ends on DNA fragments?
Allows for easier insertion into recipient cells
What characterizes a blunt-ended fragment generated by restriction endonucleases?
Aligns perfectly with complementary sequences in recipient cells
What role do vectors play in the process of DNA cloning?
Carry and replicate inserted genes in host cells
Why are restriction endonucleases crucial in creating recombinant DNA?
To generate DNA fragments with specific ends for insertion
Which enzymatic activity is characteristic of restriction endonucleases?
Recognize and cut specific sequences in DNA
What is the purpose of adding six histidines to a protein?
Enabling purification using nickel-bound beads
Which protein tag is associated with the amino acid sequence DYKDDDDK?
FLAG tag
What method is used to separate proteins according to size in gel electrophoresis?
Gel electrophoresis (SDS-PAGE)
Which protein tag allows for purification by binding to glutathione?
GST tag
Which amino acid sequence is specific to the V5 protein tag?
GKPIPNPLLGLDST
What is the detection method for Green Fluorescent Protein (GFP)?
Fluorescence detection
What is the purpose of the ShineDalgarno (SD) sequences in expression vectors?
Promote transcription initiation
In DNA cloning, what is the function of a poly-T primer?
Selecting for human mRNA
Why are eukaryotic systems like yeast preferred over bacteria for protein expression?
To enable post-translational modifications
What is the primary role of a gene clone in the creation of protein hybrids?
Creating tagged proteins
Which characteristic of restriction endonucleases distinguishes the 'many types of RNA' challenge?
Palindromic sequences
What is the impact of the lack of glycosylation on protein expression in bacteria?
Prevents post-translational modifications
How does the addition of tags affect protein purification efficiency?
Improves target protein yield
What is the function of DNA ligase in DNA cloning?
Joining DNA ends by forming phosphodiester bonds
Why are plasmids considered suitable vectors for cloning?
They possess antibiotic resistance genes
What is the purpose of an origin of replication in plasmid cloning vectors?
To ensure independent replication of the plasmid
How can a DNA fragment be inserted into a plasmid vector?
By ligating the fragment into the vector
What is the purpose of a selectable gene in a plasmid cloning vector?
To identify cells containing the vector
In cloning, what is the role of recombinant DNA technology?
Inserting DNA fragments into vectors
What is a protein domain characterized by?
Self-stabilization and independent folding
Which of the following is NOT a characteristic of a protein domain?
Dependence on the rest of the protein
What is the role of Green Fluorescent Protein (GFP) in proteins?
To allow protein detection
How do GFP-tagged proteins differ from GST-tagged proteins?
GFP allows for purification purposes
Which technique involves separating proteins based on size in gel wells?
Gel electrophoresis
Why is it essential for a protein domain to fold independently from the rest of the protein?
To maintain its compact structure
What distinguishes Green Fluorescent Protein (GFP) from Glutathione S transferase (GST) in their usage?
Purification vs. gene detection
What type of structure does a protein domain have?
$\alpha$-helix, $\beta$-sheet, or both
What is the purpose of a luciferase reporter assay in molecular biology?
To study the activity of a gene under certain conditions
In the Co-immunoprecipitation technique, what is targeted by the antibody molecules conjugated to special beads?
The protein of interest
What is the primary role of a DNA library in molecular biology?
To store DNA fragments for analysis
Which technique involves transfected cells and measures the expression level of luciferase instead of the original gene?
Luciferase reporter assay
What is the main target in Co-immunoprecipitation experiments using antibody-conjugated beads?
Protein of interest
Why would a scientist use a luciferase reporter assay instead of directly measuring the original gene's expression level?
To avoid altering the original gene
What is the purpose of the LacZ gene in the yeast two-hybrid system?
To generate a blue product in the presence of X-gal
Why are both DNA-binding domain (BD) and activation domain (AD) required to be close to each other in the yeast two-hybrid system?
To activate transcription
How is the production of recombinant proteins achieved in the cloning of hybrid proteins?
By transferring recombinant plasmids into yeast cells
What is the purpose of transferring recombinant plasmids into yeast cells in the cloning of hybrid proteins?
To identify unknown interacting proteins
Why are yeast cells grown in the presence of X-gal during the yeast two-hybrid system?
To identify interacting proteins based on color change
What distinguishes the function of DNA-binding domain (BD) and activation domain (AD) in the yeast two-hybrid system?
BD controls DNA binding, while AD enhances transcription activation.
What is the main purpose of a luciferase reporter assay in molecular biology?
Study the activity of a gene by measuring luciferase expression
In a luciferase reporter assay, what is placed upstream of the reporter gene in the plasmid?
Regulatory region of the gene
What is the specific function of Co-immunoprecipitation in molecular techniques?
Targeting proteins bound to beads
In Co-immunoprecipitation, what are antibody molecules conjugated to for targeting proteins?
Special beads
What distinguishes Firefly luciferase in a luciferase reporter assay?
Role as a reporter gene
What is the primary purpose of a DNA library?
Create bacterial clones with specific DNA pieces
What is the role of the promoter in a luciferase reporter assay using recombinant DNA techniques?
'Switch on' luciferase expression
How does Co-immunoprecipitation work in identifying proteins of interest?
'Switch on' only the protein of interest and its bound proteins
What is the role of the activation domain (AD) in the yeast two-hybrid system?
Responsible for the activation of transcription
Why are both DNA-binding domain (BD) and activation domain (AD) required to be close to each other in the yeast two-hybrid system?
To facilitate interactions between known and unknown proteins
What distinguishes the UAS in yeast cells?
Is controlled by a transcription factor
What is the significance of growing yeast cells in the presence of X-gal during the yeast two-hybrid system?
Produces a blue product upon cleavage
In the context of cloning hybrid proteins, what is cloned separately along with the unknown gene Y?
The activation domain (AD)
What is the primary purpose of using the LacZ gene in the yeast two-hybrid system?
To generate a blue product for identifying interactions
What is the main function of Cas9 in the CRISPR/Cas9 system?
Creating single or double strand breaks in DNA
How is the Cas9 nuclease directed to its target sequence in the CRISPR/Cas9 system?
By a short RNA fragment known as a guide RNA (gRNA)
What happens when a phage infects a bacterial cell in relation to the CRISPR/Cas9 system?
The bacterial cell degrades the phage DNA and integrates fragments into the CRISPR cluster
How can a gene be studied using CRISPR/Cas9 gene editing technology?
By directly altering the gene's function using Cas9
What is the specific role of the guide RNA (gRNA) in the CRISPR/Cas9 system?
Directing Cas9 to its target sequence in DNA
What is the outcome of introducing both the gRNA and Cas9 gene into human cells?
Study of gene function by mutating it
How does CRISPR/Cas9 system protect bacterial cells from phages?
By degrading the phage DNA and integrating a fragment into the CRISPR cluster
What guides the Cas9 nuclease to its target sequence in the CRISPR/Cas9 system?
Guide RNA (gRNA)
What allows for the function of a gene to be studied through gene editing mechanisms?
Mutating the gene to observe effects
How are both gRNA and Cas9 gene introduced into human cells in gene editing?
As separate genes cloned into plasmid vectors
What specifically occurs when a phage infects a bacterial cell after integration into the CRISPR/Cas9 system?
The phage DNA is transcribed into guide RNA (gRNA)
What is the role of Cas9 nuclease in gene editing using CRISPR technology?
Guiding to its target DNA sequence for cutting
How does gene editing with CRISPR technology enable replacing a mutated gene with a normal one?
Through DNA repair mechanisms using introduced DNA
'What role does gRNA play in guiding Cas9 nuclease to its target?' - What would be a correct answer?
'Directing Cas9 to cut specific DNA sequences'
Learn about DNA cloning, a technique that involves amplifying DNA segments and expressing genes in biological systems. This quiz covers the basics of forming recombinant DNA with vectors for genetic manipulation.
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