Development of effective isolation method of ES cells for analysis of differentiation.
Neuroectoderm development is a milestone of vertebrate neurogenesis. However, the molecular mechanism underlying the differentiation of neuroectoderm is still unclear, especially in mammals. ES cells co-cultured with PA6 cells can differentiate to neuroectoderm by the stromal cell-derived inducing activity method (SDIA method), but contamination of PA6 cells is an obstacle to the analysis of molecular mechanisms of differentiation. Here we describe a novel method by which differentiated ES cells are easily isolated from PA6 cells. We attempted to induce the differentiation of ES cells using paraformaldehyde-fixed PA6 cells. RT-PCR and DNA microarray analysis revealed that the background noise derived from contaminated PA6 cells disappeared when fixed PA6 cells were used. Furthermore, genes up-regulated during the differentiation of ES cells were expressed in a developing mouse embryo. Thus, our newly developed method will be very useful for identifying novel genes associated with mouse neuroectoderm development in vitro and in vivo.
Ohse
Kensuke
K
Department of Biological Science and Technology, Faculty of Industrial Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda-shi, Chiba, Japan.
Ohtsu
Masaya
M
Onoda
Fumitoshi
F
Murakami
Yasufumi
Y
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D053595:Embryonic Stem Cells / Q000166:cytology*; D020869:Gene Expression Profiling; D005819:Genetic Markers; D051379:Mice; D054258:Neural Plate / Q000166:cytology / Q000196:embryology* / Q000378:metabolism; D055495:Neurogenesis; D020411:Oligonucleotide Array Sequence Analysis
D005819:Genetic Markers