The FELIX vector system is derived from Feline Immunodeficiency Virus (FIV). FIV is a member of the lentivirus subfamily of retroviruses, and causes an immunodeficiency syndrome in cats which closely resembles that caused by HIV in humans. The genome of FIV is shown below:
The Gag polyprotein is proteolytically processed into the structural components of the virion. The Gag-Pol polyprotein is made by translational frameshifting at the Gag to Pol junction. Pol is proteolytically processed to yield the protease, reverse transcriptase, integrase and dUTPase. Vif, as in the primate lentiviruses, is believed to control virion release from lymphocytes. A, like the primate lentiviral Tat, is a transcriptional activator of the LTR; however, A is olnly a weak transactivator and does not seem to bind to any ordered secondary structure in the LTR. Rev mediates the nucleocytoplasmic export of intron-containing viral mRNAs by binding to the RRE (located in the 3' end of Envelope). The FIV envelope mediates binding and entry into lymphocytes and macrophages, among others, and is believed to use CXCR4 as its primary receptor.
II. Purpose of the FELIX Vector System
The FELIX system provides an efficient means of transferring genes via helper-free retroviral particles to non-dividing and terminally differentiated cells. Of course, the FELIX vectors are also useful for transducing dividing cells with genes of interest. FELIX vector particles carry the gene of interest into the target cell and integrate it into the target cell genome, thus the target cell is stably transduced with the gene and will retain it throughout its life.
Some applications of FELIX vectors are listed below:
Transduction in vitro of primary, non-dividing cells (e.g. CD34+ stem cells,
dendritic cells) to study their biology.
2) In vivo gene transfer for gene therapy to non-dividing cells such as hepatocytes and
CD34+ stem cells.
3) cDNA library screens in primary cells.
III. Using the FELIX System
vector system consists of three plasmids - structural, envelope, and transfer
vector - which are cotransfected into 293T cells to produce virus.
The structural construct makes Gag-Pol and Rev (some variants also make
Vif and A). Any envelope construct can be used for pseudotyping -
VSV-G tends to give the highest efficiency and broadest host range.
The transfer vector contains the viral LTRs and packaging signal. Any expression cassette (promoter and gene of interest) can be cloned into the vector. Inserts can be up to 8.5kB.
MTAs, and Plasmid Maps are available on the Felix