A genetical and biochemical investigation of the uptake of DNA by mammalian cells in culture

Lockhart, Norman Stanley (1972) A genetical and biochemical investigation of the uptake of DNA by mammalian cells in culture. PhD thesis, University of Glasgow.

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The published evidence relating to the uptake and genetic effects of purified DNA is reviewed, and the nature of polyoma pseudovirions is discussed. The characteristics of the biochemically-marked cell lines used in this work are described, and conditions for the labelling of DNA with bromodeoxyuridine and tritium are investigated. Experiments on the uptake of isolated DNA indicate that up to 5% of the DNA complement of the recipient cell is absorbed from the medium. There is a lag of 2-3 hours after addition of DNA to the medium before uptake becomes detectable, after which time uptake is much more rapid. Even at the earliest time at which donor DNA is detectable in recipient cells by autoradiography, all of it is found in the nuclei, unless DEAE-dextran is present in the medium, in which case the donor label appears in clumps in the cytoplasm. Density-labelling experiments are consistent with this, for in the presence of DEAE-dextran the donor DNA does not become associated with the density band characteristic of the recipient material. DEAE-dextran also prevents the association of donor DNA with high molecular-weight DNA in sucrose gradients, and renders the majority of DNA uptake insensitive to arabinosylcytosine. In addition to this, DNA absorbed in the presence of DEAE-dextran is lost from the cell over a period of about 48 hours after its removal from the medium. There is evidence that, even in the absence of DEAE-dextran, there is a fraction of the donor DNA which survives intact, although they are associated with' the recipient density band. Experiments are described which did not detect genetic changes in cells treated with polyoma pseudovirions or with pure DNA under a variety of conditions.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: John H Subak-Sharpe
Keywords: Biochemistry
Date of Award: 1972
Depositing User: Enlighten Team
Unique ID: glathesis:1972-73281
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 14 Jun 2019 08:56
Last Modified: 14 Jun 2019 08:56
URI: https://theses.gla.ac.uk/id/eprint/73281

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