LENTIVIRAL VECTOR PREPARATION

Procedures on lentiviral vector preparation

The success of oocytes lentiviral infections depends on the quality of lentiviral vector preparation.

CFCM is offering this service in base of the protocol set up by the laboratory of Luigi Naldini of the San Raffaele Telethon Institute for Gene Therapy.
In that way, we have not only a control on the quality of lentiviral vector preparation, but we help Users that do not have access to a P2 room.

How we proceed:

1. The User contacts CFCM describing the transgenic mouse model that want generate.
2. CFCM give suggestions on the kind of approach to follow. In the case the best solution is to use lentiviral infection, the User will sign an MTA with Luigi Naldini to receive the trasfer vector (pCCLsin.PPT.hPGK.GFP.pre) where to insert the transgene of interest. To receive the MTA, please contact Lorenza Ronfani (ronfani.lorenza@hsr.it).
3. The User performs the construct and sends to CFCM both the construct and transformed bacteria. CFCM will produce a new DNA preparation following a specific protocol necessary to produce high quality lentiviral vector.
4. CFCM produces the lentiviral vector and titers the preparation calculating the quantity of P24 protein using the p24 ELISA KIT from Perkin Elmer. In the case the transfer vector contains the GFP protein the titer will be evaluated by FACS analysis.
5. The User will receive cellular extracts from Hela transduced with different dilutions of the lentiviral production to test the protein function.
6. CFCM infects oocytes.

Here below the scheme of rules and time required for each step of your project.