Efficient isolation and gene expression profiling of small numbers of neural crest stem cells and developing Schwann cells.
Schwann cells develop from multipotent neural crest stem cells and are important for neuronal survival, maintenance of axonal integrity, and myelination. We used transgenic mice expressing green fluorescent protein in a tissue-specific manner to isolate viable, pure populations of neural crest stem cells and developing Schwann cells, which are not readily accessible by microdissection. Starting with the minute amounts of RNA obtained, a two-round amplification procedure was used to achieve reproducible DNA array hybridizations. We validated our screening procedure by comparisons with the literature and by in situ hybridization. Stage-to-stage comparisons and hierarchical clustering for neural crest and five stages of Schwann cell development suggest a wealth of candidates for genes involved in stem cell regulation and in early Schwann cell development. The combination of methods applied in this study should be generally useful for isolating and profiling other stem cell and difficult to isolate cell populations.
Buchstaller
Johanna
J
Institute of Cell Biology, Department of Biology, Swiss Federal Institute of Technology, CH-8093 Zürich, Switzerland.
Sommer
Lukas
L
Bodmer
Matthias
M
Hoffmann
Reinhard
R
Suter
Ueli
U
Mantei
Ned
N
D000818:Animals; D002452:Cell Count; D002454:Cell Differentiation / Q000502:physiology; D019070:Cell Lineage / Q000235:genetics; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D005434:Flow Cytometry; D020869:Gene Expression Profiling / Q000379:methods*; D017930:Genes, Reporter; D005865:Gestational Age; D049452:Green Fluorescent Proteins; D017403:In Situ Hybridization; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D009432:Neural Crest / Q000166:cytology*; D020411:Oligonucleotide Array Sequence Analysis; D011237:Predictive Value of Tests; D012313:RNA / Q000737:chemistry / Q000302:isolation & purification; D015203:Reproducibility of Results; D012583:Schwann Cells / Q000166:cytology* / Q000378:metabolism*; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism*
D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D012313:RNA