Targeted nucleotide exchange in Saccharomyces cerevisiae directed by short oligonucleotides containing locked nucleic acids.
Locked nucleic acids (LNAs) are novel base modifications containing a methylene bridge uniting the 2'-oxygen and the 4'-carbon. In this study, LNA-modified single-stranded molecules directed the repair of single base mutations in a yeast chromosomal gene. Using a genetic assay involving a mutant hygromycin-resistance gene, correction of point and frameshift mutations was facilitated by vectors containing an LNA residue on each terminus. Increasing the number of LNA bases on each terminus reduced the correction frequency progressively. When the LNA vector is used in combination with a phosphorothioate-modified vector (74-mer), however, a high level of gene-repair activity occurs; hence, short LNA-based vectors can augment the activity of other types of targeting vectors. These data suggest that oligonucleotides containing locked nucleic acid residues can be used to direct single nucleotide exchange reactions in vivo.
Parekh-Olmedo
Hetal
H
Department of Biological Sciences, Delaware Biotechnology Institute, University of Delaware, Newark, DE 19716, USA.
Drury
Miya
M
Kmiec
Eric B
EB
D000595:Amino Acid Sequence; D001483:Base Sequence; D004260:DNA Repair / Q000502:physiology*; D004271:DNA, Fungal / Q000235:genetics; D004277:DNA, Single-Stranded / Q000235:genetics*; D004352:Drug Resistance, Microbial / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D006921:Hygromycin B / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000737:chemistry* / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D017384:Sequence Deletion; D013873:Thionucleotides / Q000737:chemistry / Q000235:genetics
D004271:DNA, Fungal; D004277:DNA, Single-Stranded; D009841:Oligonucleotides; D013873:Thionucleotides; D006921:Hygromycin B