title: A Cre-lox approach for transient transgene expression in neural precursor cells and long-term tracking of their progeny in vitro and in vivo.
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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Clinical Neurosciences, Cambridge centre for Brain Repair, University of Cambridge, Cambridge, UK. cg355@cam.ac.uk <cg355@cam.ac.uk>
authors.forename: Cédric G
authors.initials: CG
authors.lastname: Geoffroy
authors.forename: Olivier
authors.initials: O
authors.lastname: Raineteau
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D019426:Integrases / Q000235:genetics*; D051379:Mice; D009474:Neurons / Q000166:cytology* / Q000637:transplantation; D010957:Plasmids; D051127:Scavenger Receptors, Class E / Q000235:genetics*; D013116:Spinal Cord / Q000166:cytology; D033581:Stem Cell Transplantation; D019076:Transgenes
chemical_list: C495138:Olr1 protein, mouse; D051127:Scavenger Receptors, Class E; D049452:Green Fluorescent Proteins; C045073:Cre recombinase; D019426:Integrases