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23 Cards in this Set
- Front
- Back
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what is the main goal of stable transfection?
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to permanently introduce a gene to the genome of a cell
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what sort of selectable marker is used in transfection to isolate cells that take up the gene of interest?
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antibacterial resistance
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what is the primary disadvantage of stable transfection?
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it is very time-consuming to create a stable cell line
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what is the main advantage of transient transfection?
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the procedure is quick
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does the plasmid in transient transfection replicate with the host genome? Why or why not?
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no; it does not have a eukaryotic origin of replication
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what is the common drawback of both stable and transient transfection?
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only a small percentage of cells will take up the DNA -- the process is inefficient
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in addition to a bacterial origin of replication and gene for antibacterial resistance, what must be found on gene to be introduced via transfection?
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eukaryotic enhancer
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in what form is DNA introduced in stable transfection? Through which method is the DNA introduced into genome?
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linear DNA; non-homologous end joining.
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what aspect of transfected DNA is required in stable transfection but not in transient transfection?
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resistance gene for growing eukaryotic cells
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what proteins are encoded by the gag gene in retroviruses?
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viral capsid proteins
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which enzymes are encoded by the pol gene in retroviruses?
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reverse transcriptase, integrase, and protease
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which proteins are encoded by the env gene in retroviruses?
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transmembrane and receptor-binding proteins
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what is the role of the cis-acting sequences PB and psi in the retroviral genome?
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PB is the binding sites for primers for reverse transcriptase; psi interacts with capsid proteins to allow efficient incorporation of the newly synthesized retroviral genome into viral particle
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what is the function of sequences at the end of the long terminal repeats?
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to help with insertion of the ds DNA genome into host cell chromosomes by integrase
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in a retroviral vector, which genes of the retrovirus have been replaced by the gene of interest?
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env, pol. Gag
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what is the name for the virus once its genome has integrate into that of the host?
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provirus
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what is the nature of the retroviral genome?
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ss RNA
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how are viral proteins obtained in a retroviral vector?
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they are added in trans
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what is the name for the line of cells which has been programmed to express the gag, env, and pol proteins or retroviruses?
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packaging cell line
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what is the unique advantage of having the pol, gag, and env genes on a genome apart from the retroviral genome?
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in subsequent progeny, these proteins will not be expressed and transfection will not continue indefinitely
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what is one serious problem with retroviral infections?
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they insert their genome anywhere in the cell. If the genome interrupts a host gene which encodes a protein, this can be a major problem
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what is a limitation of mouse retroviruses?
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they can only infect cells that are dividing bc genome/capsid is otherwise too large to enter nucleus.
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what is the upper limit of the size of genes of interest allowed to be transfected via retroviruses?
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8kb
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