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Questions and Answers

What is the purpose of supplementing the medium with 1-2% FCS if CHO cells are not doubling every 14-17 hours?

  • To maintain the cells in a suspended state
  • To enhance cell growth and division (correct)
  • To induce differentiation of the CHO cells
  • To inhibit cell proliferation
  • Which element in the CMV vectors is responsible for replication in bacterial cells?

  • f1 origin
  • pre-pro-trypsin leader (PPT) sequence
  • hGH polyA
  • β-lactamase gene (correct)
  • What does the SV40 replication origin contribute to in SV40 replication permissive cells?

  • Stabilization of plasmids
  • Inhibition of transcription
  • Promotion of protein secretion
  • Enhancement of DNA replication (correct)
  • Which component directs the secretion of fusion proteins into the culture medium for purification?

    <p>pre-pro-trypsin leader (PPT) sequence</p> Signup and view all the answers

    What is the function of the hCMV promoter regulatory region in the mammalian expression system?

    <p>Drives constitutive protein expression in mammalian cells</p> Signup and view all the answers

    Why are CMV vectors favorable for expressing proteins in mammalian cells?

    <p>They drive high levels of constitutive protein expression</p> Signup and view all the answers

    In the context of heterologous protein expression, what is the function of the lacI binding site in the lac operon control elements?

    <p>Inhibit T7 polymerase gene expression</p> Signup and view all the answers

    What cellular component is crucial for the production of large, complex glycosylated polypeptides in mammalian cell expression systems?

    <p>Proline</p> Signup and view all the answers

    What modification is necessary to induce constitutive expression in an un-induced state using the lac operon control elements?

    <p>Dissociation of lacI from the operator site</p> Signup and view all the answers

    Which component is responsible for the stabilization of plasmids used in bacterial cells for heterologous protein expression?

    <p>Lac Repressor (lacI)</p> Signup and view all the answers

    What characteristic makes Chinese Hamster Overy (CHO) cells a preferred host cell for mammalian expression systems?

    <p>They can be grown as a suspension in bioreactors</p> Signup and view all the answers

    Which element of the lac operon control system directly affects the expression of the gene of interest in bacterial cells?

    <p>Lac Repressor (lacI)</p> Signup and view all the answers

    What is the primary control point of the CMV promoter?

    <p>Enhancer Sequence</p> Signup and view all the answers

    Which cell type shows a relatively high level of gene expression with the CMV promoter?

    <p>CHO cells</p> Signup and view all the answers

    What factor is crucial for the tight control of growth conditions when using the CMV enhancer sequence?

    <p>Cellular Differentiation</p> Signup and view all the answers

    Which transcription factor has a strong preference for the CMV enhancer sequence?

    <p>CREB</p> Signup and view all the answers

    What type of cells show low levels of gene expression with the CMV promoter?

    <p>Stem cells</p> Signup and view all the answers

    What is one disadvantage associated with the CMV enhancer sequence?

    <p>Tight control of growth conditions required</p> Signup and view all the answers

    What is one potential issue with using the lac operon control elements for heterologous protein expression?

    <p>Leaky expression leading to plasmid instability</p> Signup and view all the answers

    What is a characteristic of the T7 RNA Polymerase in the context of heterologous protein expression?

    <p>Under full induction, it can result in the desired protein being over 50% of total cell protein</p> Signup and view all the answers

    What is the main role of the lac repressor in the lac operon system?

    <p>To bind to the operator and prevent transcription in the absence of inducers</p> Signup and view all the answers

    Why is it important to remove the lacZYA genes when using the lac operon for heterologous protein expression?

    <p>To prevent interference with translation of the gene of interest</p> Signup and view all the answers

    Which statement accurately describes the role of Galactoside permease (lacY) in the lac operon system?

    <p>Allows lactose uptake from media into bacterial cells</p> Signup and view all the answers

    Why is a more tightly controlled promoter-expression-cassette needed when using the lac operon for heterologous protein expression?

    <p>To prevent target gene overexpression and cellular damage</p> Signup and view all the answers

    What is the main purpose of using different restriction enzymes to cut the donor and receiver DNA when cloning a gene?

    <p>To prevent re-ligation of DNA fragments.</p> Signup and view all the answers

    In the context of biotechnology, what is the role of T7 lysozyme in host cells like E. coli?

    <p>Breaks down T7 Polymerase before induction.</p> Signup and view all the answers

    Which type of promoter is commonly used in eukaryotic expression vectors for strong gene expression?

    <p>CMV promoter</p> Signup and view all the answers

    Why is orientation important when cloning genes using different restriction enzymes?

    <p>To ensure correct expression of the gene.</p> Signup and view all the answers

    What is one advantage of using mammalian cells as an expression host over bacterial or yeast cells?

    <p>Easier post-transcriptional modifications.</p> Signup and view all the answers

    Why is it necessary to choose the appropriate expression format at the beginning of cloning work in biotechnology?

    <p>To ensure proper folding and modification of proteins.</p> Signup and view all the answers

    What are the major disadvantages of using inducible expression systems in biotechnology?

    <p>off-target effects and high basal expression level</p> Signup and view all the answers

    Why are stable inducible mammalian expression vectors considered the 'holy grail' for industrial recombinant protein expression?

    <p>They allow for humanized product from a stable cell line that can be controllably induced without inducing pleiotropic effects.</p> Signup and view all the answers

    What is a concern associated with stable integration of genes into the genome in biotechnology?

    <p>Down-regulation of the promoter through methylation/de-acetylation</p> Signup and view all the answers

    What are some examples of inducible systems used in biotechnology and their respective inducers?

    <p>Temperature shift (37-42°C), heavy metals (zinc, copper, cadmium), and steroids (glucocorticoids)</p> Signup and view all the answers

    Why is serum batch-to-batch variability a concern in modulating enhancers in biotechnology?

    <p>It can lead to unpredictable changes in enhancer activity and affect the overall expression of the target gene.</p> Signup and view all the answers

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