A DNA transposon-based approach to functional screening in neural stem cells.
We describe the use of DNA transposons as tools for carrying out functional screenings in murine embryonic stem (ES) cell-derived neural stem (NS) cells. NS cells are a new type of stem cells featuring radial glial properties, that undergoes symmetric cell division for an indefinite number of passages, expanding as a monolayer. In this model, the previously unreported Sleeping Beauty transposase M3A achieves an optimal blend of clone generation efficiency and low redundancy of integrations per clone, compared to the SB100X Sleeping Beauty variant and to the piggyBac transposon. The technology described here makes it possible to randomly trap genes in the NS cell genome and modify their expression or tag them with fluorescent markers and selectable genes, allowing recombinant cells to be isolated and expanded clonally. This approach will facilitate the identification of novel determinants of stem cell biology and neural cell fate specification in NS cells.
Albieri
Ilaria
I
Division of Neuroscience, San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milano, Italy.
Onorati
Marco
M
Calabrese
Giovanna
G
Moiana
Alessia
A
Biasci
Daniele
D
Badaloni
Aurora
A
Camnasio
Stefano
S
Spiliotopoulos
Dimitrios
D
Ivics
Zoltán
Z
Cattaneo
Elena
E
Consalez
G Giacomo
GG
D000818:Animals; D002478:Cells, Cultured; D003198:Computer Simulation; D004251:DNA Transposable Elements / Q000235:genetics*; D006801:Humans; D051379:Mice; D008957:Models, Genetic; D016254:Mutagenesis, Insertional / Q000379:methods*; D009355:Neomycin; D058953:Neural Stem Cells / Q000502:physiology*; D019895:Transposases / Q000235:genetics* / Q000378:metabolism
D004251:DNA Transposable Elements; D019895:Transposases; C494471:sleeping beauty transposase, human; D009355:Neomycin