The detection of oligodeoxynucleotide molecules following uptake into mammalian cells.
A mixed phosphodiester:phosphorothioate oligodeoxynucleotide was used in uptake studies with T15 mouse fibroblast cells. The presence of full-length unlabeled oligomers was identified in both cytoplasmic and nuclear extracts by a method involving gel electrophoresis and electroblotting followed by hybridization with a complementary radiolabeled probe. Detection did not depend on the presence of a label on the oligomer with the potential for its removal, often a problem in other studies. The fate of the oligonucleotides could then be followed with time for at least 3 days. This method of detection should be applicable to studies of nuclease resistance and uptake characteristics of newly developed oligonucleotide analogues.
Hawley
P
P
University of East Anglia, School of Biological Sciences, Norwich, UK.
Gibson
I
I
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000097:blood / Q000378:metabolism*; D013873:Thionucleotides / Q000097:blood / Q000378:metabolism*
D009838:Oligodeoxyribonucleotides; D013873:Thionucleotides