Abstract
Background
Gene transfer is a potential treatment modality of genetic disease. Efficient, practical methods of DNA transfection are currently under investigation.
Materials and Methods
A β-galactosidase reporter plasmid interacted electrostatically with his-tones, poly-l-Lys, poly-l-Arg, and a combination of poly-l-Lys and poly-l-Arg. This complex was then used to transfect COS-7 cells, β-galactosidase activity was quantified and used to compare the efficiency of gene transfection in vitro. A comparison was also made of DNA transfection with the most active histone subclass, i.e., histone H2A, in the absence and presence of an anionic liposome.
Results
There was a marked increase in DNA transfection in the presence of histone H2A when compared with the control, whereas each of the other histones and polycations showed little, if any, effect. The extent of activation depends strongly on the DNA/histone ratio and is also a function of the molarity of the final Tris-acetate, pH 8, solution. The anionic liposomes used demonstrated an inhibitory effect.
Conclusions
Histone H2A significantly enhances in vitro DNA transfection whereas other histones and anionic liposomes do not. A study of the difference between histone H2A and other histone subclasses may serve to clarify some of the mechanisms and the essential components of efficient gene delivery.
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Acknowledgements
Drs. P. L. Feigner and O. Zelphati (Vical, Inc., San Diego, CA) are gratefully acknowledged for numerous helpful discussions. We thank Dr. Timo L. M. ten Hagen (Rotterdam, The Netherlands) for his gift of PC/PS liposomes that were used in preliminary experiments. Susan Sackmann is gratefully acknowledged for skillful technical assistance. D. Balicki is a recipient of The Medical Research Council of Canada’s Clinician-Scientist Award. This is manuscript 11055-MEM from The Scripps Research Institute. This work was supported by National Institutes of Health grant RR00833 and the Stein Endowment Fund.
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Balicki, D., Beutler, E. Histone H2A Significantly Enhances In Vitro DNA Transfection. Mol Med 3, 782–787 (1997). https://doi.org/10.1007/BF03401715
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DOI: https://doi.org/10.1007/BF03401715