Simultaneous targeted exchange of two nucleotides by single-stranded oligonucleotides clusters within a region of about fourteen nucleotides.
BACKGROUND
Transfection of cells with gene-specific, single-stranded oligonucleotides can induce the targeted exchange of one or two nucleotides in the targeted gene. To characterize the features of the DNA-repair mechanisms involved, we examined the maximal distance for the simultaneous exchange of two nucleotides by a single-stranded oligonucleotide. The chosen experimental system was the correction of a hprt-point mutation in a hamster cell line, the generation of an additional nucleotide exchange at a variable distance from the first exchange position and the investigation of the rate of simultaneous nucleotide exchanges.


RESULTS
The smaller the distance between the two exchange positions, the higher was the probability of a simultaneous exchange. The detected simultaneous nucleotide exchanges were found to cluster in a region of about fourteen nucleotides upstream and downstream from the first exchange position.


CONCLUSIONS
We suggest that the mechanism involved in the repair of the targeted DNA strand utilizes only a short sequence of the single-stranded oligonucleotide, which may be physically incorporated into the DNA or be used as a matrix for a repair process.
Hegele
Heike
H
Institute of Human Genetics, University of Ulm, D 89070 Ulm, Germany. Heike.Hegele@t-online.de
Wuepping
Matthias
M
Ref
Caroline
C
Kenner
Oliver
O
Kaufmann
Dieter
D
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D017931:DNA Primers / Q000235:genetics; D004260:DNA Repair / Q000235:genetics / Q000502:physiology*; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D017931:DNA Primers; D004277:DNA, Single-Stranded; D009838:Oligodeoxyribonucleotides; D007041:Hypoxanthine Phosphoribosyltransferase