Podcast
Questions and Answers
Which of the following techniques is LEAST likely to be used for the direct introduction of recombinant DNA into a host cell?
Which of the following techniques is LEAST likely to be used for the direct introduction of recombinant DNA into a host cell?
- Gene gun (biolistics)
- Electroporation
- Reverse transcription (correct)
- Microinjection
A scientist is trying to produce a recombinant protein in E. coli. They have successfully transformed the bacteria with a plasmid containing the gene of interest under the control of a strong promoter. However, they observe very little protein production. Which of the following is the LEAST likely reason for this?
A scientist is trying to produce a recombinant protein in E. coli. They have successfully transformed the bacteria with a plasmid containing the gene of interest under the control of a strong promoter. However, they observe very little protein production. Which of the following is the LEAST likely reason for this?
- The *E. coli* strain lacks the necessary transcription factors to activate the promoter on the plasmid. (correct)
- The recombinant protein is being degraded by intracellular proteases.
- The protein is insoluble and forming inclusion bodies.
- The mRNA is not being efficiently translated due to codon bias.
In a typical PCR, which of the following steps involves the binding of primers to the single-stranded DNA template?
In a typical PCR, which of the following steps involves the binding of primers to the single-stranded DNA template?
- Annealing (correct)
- Ligation
- Elongation
- Denaturation
Which of the following is NOT a typical component of a plasmid vector used in genetic engineering?
Which of the following is NOT a typical component of a plasmid vector used in genetic engineering?
After creating a recombinant plasmid, you need to introduce it into bacterial cells. Which method relies on using brief electrical pulses to create temporary pores in the cell membrane, allowing the plasmid to enter?
After creating a recombinant plasmid, you need to introduce it into bacterial cells. Which method relies on using brief electrical pulses to create temporary pores in the cell membrane, allowing the plasmid to enter?
Consider a scenario where a researcher aims to clone a specific gene into a plasmid vector. After digestion of both the gene and the vector with the same restriction enzyme, the researcher attempts ligation but finds very few colonies after transformation. What could be the MOST probable reason for this outcome?
Consider a scenario where a researcher aims to clone a specific gene into a plasmid vector. After digestion of both the gene and the vector with the same restriction enzyme, the researcher attempts ligation but finds very few colonies after transformation. What could be the MOST probable reason for this outcome?
In the context of downstream processing in biotechnology, what does 'formulation' primarily refer to?
In the context of downstream processing in biotechnology, what does 'formulation' primarily refer to?
Which of the following is the MOST direct application of using restriction enzymes in recombinant DNA technology?
Which of the following is the MOST direct application of using restriction enzymes in recombinant DNA technology?
A scientist is working on expressing a eukaryotic gene in a bacterial system. They include an inducible promoter in their construct. What is the primary reason for using an inducible promoter rather than a constitutive promoter?
A scientist is working on expressing a eukaryotic gene in a bacterial system. They include an inducible promoter in their construct. What is the primary reason for using an inducible promoter rather than a constitutive promoter?
What is the primary purpose of including a selectable marker gene, like antibiotic resistance, in a plasmid vector?
What is the primary purpose of including a selectable marker gene, like antibiotic resistance, in a plasmid vector?
Flashcards
Biotechnology
Biotechnology
The use of living organisms or biological systems to develop or make products.
Genetic Engineering
Genetic Engineering
The process of manipulating an organism's genes using biotechnology.
Recombinant DNA
Recombinant DNA
Combining DNA from different sources to create new genetic combinations.
Restriction Enzymes
Restriction Enzymes
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Plasmids
Plasmids
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Transformation
Transformation
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Cell Culture
Cell Culture
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Downstream Processing
Downstream Processing
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PCR (Polymerase Chain Reaction)
PCR (Polymerase Chain Reaction)
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Gel Electrophoresis
Gel Electrophoresis
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Study Notes
Biotechnology Overview
- Biotechnology involves using living organisms, cells, or their components to develop technologies and products
- It leverages biological systems to create advancements in various fields
- Includes genetic engineering, tissue culture, and other techniques
- Plays a crucial role in medicine, agriculture, and industry
Principles of Biotechnology
- Genetic engineering involves altering an organism's genetic material to achieve desired traits
- Aseptic environment maintenance is essential to prevent contamination during bioprocessing
- Modifying genetic material to introduce it into other organisms is a key aspect
- Biotechnology harnesses natural biological processes for human benefit
First Recombinant DNA
- Stanley Cohen and Herbert Boyer conducted experiments
- They cut DNA from the Salmonella typhimurium plasmid using restriction enzymes
- They then linked this DNA to a plasmid
- The recombinant plasmid was introduced into E. coli, demonstrating gene cloning
Steps in Genetic Engineering
- Identifying a DNA sequence with a desirable gene is the first step
- The gene must then be introduced into a host organism
Steps of Biotechnology
- Involves obtaining the desired product on a large scale
- Downstream processing is a crucial step for purifying and formulating the product
Identification and Isolation
- Identifying a DNA sequence with a desirable gene is a key step in recombinant DNA technology
- Isolation of the desired gene is also essential for further manipulation
Fragmentation
- Restriction enzymes are used to cut DNA into fragments
- These enzymes recognize specific sequences and cut the DNA at those sites
- This process is crucial for creating DNA fragments for cloning and other applications
Separation
- Gel electrophoresis is commonly used to separate DNA fragments based on size
- DNA fragments move through the gel matrix when an electric field is applied
- Smaller fragments migrate faster, resulting in separation by size
Amplification
- Polymerase Chain Reaction (PCR) is a technique to amplify specific DNA sequences
- PCR involves repeated cycles of denaturation, annealing, and extension
- This results in exponential amplification of the target DNA sequence
Ligation
- DNA ligase is an enzyme that joins DNA fragments together
- This process is essential for creating recombinant DNA molecules
- Ligase forms phosphodiester bonds between the fragments
Transformation
- The introduction of recombinant DNA into a host cell
- Common methods include electroporation and heat shock
- Successful transformation results in the host cell carrying the desired gene
Culture
- Host cells containing recombinant DNA are cultured to produce the desired product
- Large-scale cultures are grown in bioreactors
- Culture conditions are optimized for maximum product yield
Downstream Processing
- Involves separation, purification, and formulation of the desired product
- Ensures the final product is of high quality and purity
- Includes clinical trials and quality control for pharmaceuticals
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