A Cre-lox approach for transient transgene expression in neural precursor cells and long-term tracking of their progeny in vitro and in vivo

Authors:
Geoffroy CG, Raineteau O
In:
Source: BMC Dev Biol
Publication Date: (2007)
Issue: 7: 45
Research Area:
Neurobiology
Stem Cells
Cells used in publication:
Neural stem cell (NSC), mouse
Species: mouse
Tissue Origin: brain
Platform:
Nucleofectorâ„¢ I/II/2b
Abstract
BACKGROUND: Neural precursor cells (NPCs) can be isolated from various regions of the postnatal central nervous system (CNS). Manipulation of gene expression in these cells offers a promising strategy to manipulate their fate both in vitro and in vivo. In this study, we developed a technique that allows the transient manipulation of single/multiple gene expression in NPCs in vitro, and the long-term tracking of their progeny both in vitro and in vivo. RESULTS: In order to combine the advantages of transient transfection with the long-term tracking of the transfected cells progeny, we developed a new approach based on the cre-lox technology. We first established a fast and reliable protocol to isolate and culture NPCs as monolayer, from the spinal cord of neonatal transgenic Rosa26-YFP cre-reporter mice. These cells could be reliably transfected with single/multiple plasmids by nucleofection. Nucleofection with mono- or bicistronic plasmids containing the Cre recombinase gene resulted in efficient recombination and the long-term expression of the YFP-reporter gene. The transient cre-expression was non-toxic for the transfected cells and did not alter their intrinsic properties. Finally, we demonstrated that cre-transfected cells could be transplanted into the adult brain, where they maintained YFP expression permitting long-term tracking of their migration and differentiation. CONCLUSION: This approach allows single/multiple genes to be manipulated in NPCs, while at the same time allowing long-term tracking of the transfected cells progeny to be analyzed both in vitro and in vivo.