SV40 ori-gebied

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SV40 ori-gebied

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Important characteristics
SV40 transducing vectors packageable and
replicating - replacing the late or early
DNA - double infection, with a helper virus
(ts) to produce viral particles - Late
replacement vectors with a tsA mutant (in
T-antigen gene) as helper - Early replacement
vectors with a tsB mutant of the VP1 gene as
helper - infected cells are recognized by
plaque formation gt disadvantage the plaques
are a mixture of reconbinant and helper virus -
construction of COS cells (Gluzman) defective
SV40 DNA integrated in CV1 cell genome (mutant
ORI by 6-bp deletion T-antigen remains
active) - Early replacement vector
transfection of COS cells no helper virus
needed gt plaques contain recombinant virus
only

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Important characteristics
SV40 plasmid vectors replicating,
non-packageable - in COS cells, after
replacing late early DNA the DNA
replicates to high copy number hence it turns
out that only ORI is required to maintain the DNA
e.g. pSV2 (as long as T-antigen is present in
trans) - obviously, no plaques are produced
(no viral coat protein is available) - the
T-antigen gene can also be provided on the vector
(modular construct) e.g. pSV3 - these vectors
are ordinarily E. coli mammalian cell shuttle
vectors SV40 passive vectors non-replicating,
non-packageable - usually shuttle vectors
not restricted to monkey cells - permanently
transformed cells, with integration of the DNA in
the genome gt form colonies (foci) under
selection pressure

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