Techniques of Chromosome-Mediated Gene Transfer
Functional genes can be transferred to eukaryotic cells by incubation with isolated metaphase chromosomes. Uptake occurs by phagocytosis and most of the ingested chromosomal DNA is degraded to small inactive fragments. The frequency of transfer of any selected marker ranges from about 10-5 to 10-7. Hence, the isolation of transformants requires a sensitive selection system. A free functional chromosome fragment can be retained in the progeny of a transformant for many generations after uptake if selection is maintained. In the absence of selection, the transferred genes are lost from progeny at a rate of about 2–10% per generation (Klobutcher and Ruddle, 1979). Stabilization of transferred genes occurs at a much lower frequency and results from integration of the fragment into nonhomologous sites on recipient chromosomes (Fournier and Ruddle, 1977). The size of the transferred fragment is usually considerably decreased during the process of integration (Klobutcher and Ruddle, 1979; Olsen et al., 1981).
KeywordsSucrose DMSO Glycine Sedimentation Fractionation
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- McBride, O. W., and Ozer, H. L., 1973, Gene transfer with purified mammalian chromosomes, in: Possible Episomes in Eukaryotes; Le petit Colloquia on Biology and Medicine, Volume 4 (L. G. Silvestri, ed.), North-Holland, Amsterdam, pp. 255–267.Google Scholar