High-throughput screening of gene function in stem cells using clonal microarrays.
We describe a microarray-based approach for the high-throughput screening of gene function in stem cells and demonstrate the potential of this method by growing and isolating clonal populations of both adult and embryonic neural stem cells. Clonal microarrays are constructed by seeding a population of cells at clonal density on micropatterned surfaces generated using soft lithographic microfabrication techniques. Clones of interest can be isolated after assaying in parallel for various cellular processes and functions, including proliferation, signal transduction, and differentiation. We demonstrate the compatibility of the technique with both gain- and loss-of-function studies using cell populations infected with cDNA libraries or DNA constructs that induce RNA interference. The infection of cells with a library prior to seeding and the compact but isolated growth of clonal cell populations will facilitate the screening of large libraries in a wide variety of mammalian cells, including those that are difficult to transfect by conventional methods.
Ashton
Randolph S
RS
The Howard P. Isermann Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, New York 12180, USA.
Peltier
Joseph
J
Fasano
Christopher A
CA
O'Neill
Analeah
A
Leonard
Joshua
J
Temple
Sally
S
Schaffer
David V
DV
Kane
Ravi S
RS
D000818:Animals; D002999:Clone Cells; D005260:Female; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D051381:Rats; D011916:Rats, Inbred F344; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology*