Spontaneous uptake of biologically active recombinant DNA by mammalian cells via a selected DNA segment.
DNA can be internalized by mammalian cells without taking advantage of helper reagents. Here, we ask whether the spontaneous cellular uptake of double-stranded DNA (dsDNA) occurs in a biologically significant and sequence-dependent way. We describe a combinatorial approach to search for dsDNA sequence segments that are preferentially internalized. A selected dsDNA species was identified and covalently linked to a luciferase expression cassette. The increased apparent cellular uptake of long-chain recombinant DNA accompanied by an increased apparent expression of luciferase provides strong evidence for the view that (i) naked long-chain dsDNA can be taken up spontaneously by mammalian cells, (ii) specific sequences substantially increase this process, and (iii) dsDNA is transported into the nucleus of cells in a bioactive form. Experimental evidence indicates a tissue- or cell-type specificity for this process. This work indicates that, in principle, specific nucleotide sequences can facilitate the introduction of naked dsDNA into target cells of interest, thereby improving existing vector systems and providing a new methodology to study DNA uptake by mammalian cells. The cellular uptake of biologically active genetic material in vivo occurs to be conceivable.
Lehmann
M J
MJ
Institut für Molekulare Medizin, Universität zu Lübeck, Ratzeburger Allee 160, Lübeck, Germany.
Sczakiel
G
G
D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured / Q000378:metabolism*; D015316:Genetic Therapy / Q000379:methods*; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000032:analysis / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D011994:Recombinant Proteins; D008156:Luciferases