Structural aspects of histone H1-DNA complexes and their relation to transfection efficiency.
During transfection, polycation-DNA complexes are normally diluted by the transfection medium, which often contains salt in the physiological concentration range and serum. It is not exactly known to what extent this dilution step influences the properties of the complexes, which in turn influence the transfection efficiency. In order to gain more insight into the size-structure-transfection activity relationship, we prepared histone H1-DNA complexes in NaCl solutions at various concentrations known to determine the size and structure of the resulting complexes. We characterized the complexes by physicochemical methods. Fluorescence correlation spectroscopy enabled relative measurements of complex sizes even under physiological conditions. The different appearances of the complexes were correlated with their transfection efficiency. When transfection was performed by dilution of the complexes in cell-cultivation media, the initial structure of H1-DNA complexes preformed under distinct salt conditions had no significant influence on the transfection efficiency. The dilution of the preformed complexes with cell-cultivation medium resulted in re-formation and aggregation of the complexes. The addition of the complexes to the cells without cell-cultivation medium, however, showed a direct correlation between the size of the complexes and the transfection efficiency (correlation coefficient 0.91). Small complexes did not contribute to the transfection.
Haberland
Annekathrin
A
Pharmacology and Toxicology, Institute of Pharmacy, Free University of Berlin, Königin-Luise-Strasse 2+4, D-14195 Berlin, Germany. ahaber@zedat.fu-berlin.de
Cartier
Régis
R
Heuer
Diane
D
Zaitsev
Sergei
S
Paulke
Bernd-R
BR
Schäfer-Korting
Monika
M
Böttger
Michael
M
D000818:Animals; D002417:Cattle; D002461:Cell Line, Transformed; D004247:DNA / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D003850:Deoxyribonuclease I; D006657:Histones / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D012965:Sodium Chloride; D013050:Spectrometry, Fluorescence; D013950:Thymus Gland / Q000378:metabolism; D014162:Transfection / Q000379:methods
D006657:Histones; D012965:Sodium Chloride; D004247:DNA; D003850:Deoxyribonuclease I