Microarray analysis of embryonic stem cells and differentiated embryoid bodies.
By altering the cellular microenvironment and culture media composition, embryonic stem cells (ESCs) can be induced to differentiate in vitro into somatic cell types from the three primitive germ layers. ESC differentiation is regulated by an intricate series of signaling events that result in their transcriptional reprogramming, asymmetric cell division, and differentiation. Using various pharmacological agents and/or genetic manipulations, one can drive and enrich ESC differentiation to specific cell lineages. Identifying the transcriptional fingerprint during ESC differentiation could yield novel targets for genetic or pharmacological regulation to increase the yield of desirable cell types. We discuss here how to culture undifferentiated mouse ESCs (E14 line from 129/Ola) and generate embryoid bodies (EBs). We also discuss in detail how to prepare Affymetrix samples, how to hybridize and scan arrays, and how to quality control data and generate signal values and permutation based P-values.
Zambon
Alexander C
AC
Department of Pharmacology, University of California, San Diego, La Jolla, CA, USA.
Barker
Christopher S
CS
D000818:Animals; D002449:Cell Aggregation; D002454:Cell Differentiation; D002460:Cell Line; D018076:DNA, Complementary / Q000096:biosynthesis / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D009693:Nucleic Acid Hybridization; D011786:Quality Control; D012313:RNA / Q000737:chemistry / Q000302:isolation & purification; D013194:Staining and Labeling; D046888:Tissue Array Analysis / Q000379:methods*
D018076:DNA, Complementary; D012313:RNA