Lentivirus production:
IMPORTANT: The biosafety office at your institution must be notified prior to use
of this system for permission and for further institution-specific instructions. BL2
conditions should be used at all times when handling the virus. All decontamination
steps should be performed using 70% ethanol/1% SDS. Gloves should be worn at
all times when handling lentiviral preparations, transfected cells or the combined
transfection reagent. Just remember that although this virus has been significantly
modified for biosafety, it derived from HIV and with a VSV pseudotype human cells
can be infected even if they are not dividing. That said, the following modifications
have been made to prevent viral replication.
1. Packaging vector lacks both LTRs and has no viral packaging signal (ψ)
2. The following viral genes have been deleted from the packaging vector:
env, tat, rev, vpr, vpu, vif and nef.
3. Rev is supplied in trans on a different vector (RSV-Rev).
4. The vector expressing the packaged viral genome has a self-inactivating
LTR (TATA box deletion) and expresses no viral gene products.
5. Envelope, in this case VSVG, is expressed on a separate vector.
For more information please refer to the following papers.
Packaging vectors pMDLg/pRRE, CMV-VSVG and RSV-Rev):
Dull et al., A Third-Generation Lentivirus Vector with a Conditional
Packaging System. J. Virol. 1998 72(11): 8463-8472.
Naldini L, Blomer U, Gallay P, Ory D, Mulligan R, Gage FH, Verma IM,
Trono D. In vivo gene delivery and stable transduction of nondividing
cells by a lentiviral vector. Science. 1996 Apr 12;272(5259):263-7.
Self inactivating LTR:
Miyoshi H, Blomer U, Takahashi M, Gage FH, Verma IM. Development of a
self-inactivating lentivirus vector. J Virol. 1998 Oct;72(10):8150-7.
