Polyethylene glycol (PEG), a highly hydrated polymer, was shown to induce cell fusion in the 1970s (1,2). Since then, this polymer has been widely used to mediate cell fusion for production of somatic cell hybrids that are invaluable tools in gene mapping, gene expression, the analysis of gene function, and the production of vaccines and antibodies (3-5). PEG-mediated cell fusion has also been used for nuclear transfer in mammalian cloning (6). PEG induces cell agglutination and cell-to-cell contact, leading to subsequent cell fusion. However, the detailed mechanisms underlying PEG-mediated cell fusion are not known. Model systems were used to define the molecular details of the fusion process (7). These studies indicate that small perturbations in lipid packing within contacting bilayer leaflets are necessary and probably sufficient to

From: Methods in Molecular Biology, vol. 325: Nuclear Reprogramming: Methods and Protocols Edited by: S. Pells © Humana Press Inc., Totowa, NJ

promote membrane fusion. In this chapter, we describe a basic protocol of PEG-mediated cell fusion for the production of somatic cell hybrids. The main procedures of the protocol include preparation of donor and recipient cells, cell fusion, hybrid selection and cloning, and characterization of hybrid cells as summarized in Fig 1A. Fusion can be performed between adherent and suspension cells or between adherent cells or suspension cells. Either whole cells or microcells can be used as donors to fuse with recipient cells. Microcell fusion is particularly useful for transfer of a single or a limited number of chromosomes between various types of cells. Using this method, we have successfully introduced mammalian minicchromosomes into a variety of vertebrate cells (8,9). The technique described here can be adapted for uses in other cell fusion involved research. This protocol, in principle, provides guidelines for further development of PEG mediated cell fusion technology.

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