Lentivirus

  • Infection of dividing and non-dividing cells
  • Long-lasting transgene expression, genome-integrating
  • 10 kb DNA uptake capacity
  • 2 ssRNA genome
  • 100 nm capsid size
  • Security level S2

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Lentiviral vectors and lentivirus production

All vectors are based on the lentiviral shuttle vector FUGW by Lois et al. 2002. In this base vector, the promoter was replaced and a multiple cloning site inserted to facilitate cloning of desired transgenes. Currently we use the following promoters: CMV, CamKII, hSynapsinI, GFAP or hUbiquitin C.

In a second type of LV vectors we employed different reporter genes together with a self-cleaving 2A peptide from porcine teschovirus-1 (P2A) followed by the MCS. This allows us the bicistronic expression of reportergene and transgene making it possible to carefully characterize virus efficiency.

The third set of viral vectors utilizes the combination of two different Lox sites. The introduction of a reporter gene plus a reverse oriented reporter gene in conjunction with P2A and MSC make these vectors suitable for cell-type specific expression of the transgene in Cre transgenic animals.

For reducing endogenous mRNAs levels (knockdown) we use a viral vector which contains the human RNA polymerase III promoter U6 for driving the transciption of specific shRNAs. A second promoter (hUbiquitin C or hSynapsin1) drives the expression of different reporter genes.

All viruses are generated using two "helper" plasmids that provide packaging and envelope proteins (packaging plasmid pCMV-dR8.9 and envelope plasmid pCMV-VCV-G . HEK293 cells are transfected with the plasmids. After 2 to 3 days of incubation, the virus supernatant is harvested and purified.

Costs - lentiviral particle production

Service

Costs

Discription

App. 10^6-10^7 particles / ml

   

Small-scale production

230€

Production in HEK293 cells, volume 10ml

Small-scale production concentrated

250€

Production in HEK293 cells, concentrated with Amicon Filters, volume 500µl

Medium-scale production

280€

Production in HEK293 cells, volume 20ml

Medium-scale production concentrated

310€

Production in HEK293 cells, concentrated with Amicon Filters, volume 1ml

large-scale production

380€

Production in HEK293 cells, volume 40ml

large-scale production concentrated

420€

Production in HEK293 cells, concentrated with Amicon Filters, volume 2ml

App. 10^9 Particles / ml

   

Small-scale production

550€

Production in HEK293 cells, Purification by ultracentrifugation, volume app. 100µl

Large-scale production

850€

Production in HEK293 cells, Purification by ultracentrifugation, volume app. 200µl