title: Determining cell division symmetry through the dissection of dividing cells using single-cell expression analysis.
abstract: Symmetric cell divisions give rise to two sister cells that are identical to each other, whereas asymmetric divisions produce two sister cells with distinctive phenotypes. Although cell division symmetry is usually determined on the basis of a few markers or biological functions, the overall similarity between sister cells has not been thoroughly examined at a molecular level. Here we provide a protocol to separate sister embryonic stem cells (ESCs) and to conduct multiplexed gene expression analyses at the single-cell level by using 48 ESC genes. The procedure includes the dissection of dividing, paired sister cells by micromanipulation, followed by cell lysis, reverse transcription, gene-specific cDNA amplification and multiplexed quantitative PCR analyses. This protocol can be completed in 10 d, and it can be readily adapted to other cell types that are able to grow in suspension culture.
authors: Haemato-Oncology Research Unit, Division of Molecular Pathology, Division of Cancer Biology, The Institute of Cancer Research (ICR), Sutton, UK.
authors: Lukasz
authors: L
authors: Jasnos
authors: Haemato-Oncology Research Unit, Division of Molecular Pathology, Division of Cancer Biology, The Institute of Cancer Research (ICR), Sutton, UK.
authors: Tomoyuki
authors: T
authors: Sawado
mesh_terms: D000818:Animals; D002455:Cell Division / Q000502:physiology*; D002469:Cell Separation / Q000379:methods*; D018076:DNA, Complementary / Q000235:genetics; D053595:Embryonic Stem Cells / Q000378:metabolism / Q000502:physiology*; D020869:Gene Expression Profiling; D051379:Mice; D008846:Micromanipulation / Q000379:methods; D060885:Multiplex Polymerase Chain Reaction; D048348:Reverse Transcription / Q000502:physiology
chemical_list: D018076:DNA, Complementary