Effects of target sequence and sense versus anti-sense strands on gene correction with single-stranded DNA fragments.
The correction of an inactivated hygromycin resistance and enhanced green fluorescent protein (Hyg-EGFP) fusion gene by a several hundred-base single-stranded (ss) DNA fragment has been reported. In this study, the effectiveness of this type of gene correction was examined for various positions in the rpsL gene. Sense and anti-sense ssDNA fragments were prepared, and the gene correction efficiencies were determined by co-introduction of the target plasmid containing the gene with the ssDNA fragments. The gene correction efficiency varied (0.8-9.3%), depending on target positions and sense/anti-sense strands. Sense ssDNA fragments corrected the target gene with equal or higher efficiencies as compared to their anti-sense counterparts. The target positions corrected with high efficiency by the sense fragments also tended to be corrected efficiently by the anti-sense fragments. These results suggest that the sense ssDNA fragments are useful for the correction of mutated genes. The variation in the correction efficiency may depend on the sequence of the target position in double-stranded DNA.
Kamiya
Hiroyuki
H
Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-ku, Sapporo, Japan. hirokam@pharm.hokudai.ac.jp
Uchiyama
Masayuki
M
Nakatsu
Yoshimichi
Y
Tsuzuki
Teruhisa
T
Harashima
Hideyoshi
H
D000818:Animals; D019677:Artificial Gene Fusion; D001483:Base Sequence; D016466:CHO Cells; D002934:Cinnamates / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D016373:DNA, Antisense / Q000737:chemistry / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics*; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D005818:Genetic Engineering; D015316:Genetic Therapy; D049452:Green Fluorescent Proteins / Q000235:genetics; D006921:Hygromycin B / Q000031:analogs & derivatives / Q000494:pharmacology; D009154:Mutation; D000091243:Ribosomal Protein S9; D012269:Ribosomal Proteins / Q000235:genetics; D014162:Transfection
D002934:Cinnamates; D016373:DNA, Antisense; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D029968:Escherichia coli Proteins; D000091243:Ribosomal Protein S9; D012269:Ribosomal Proteins; C000717190:RpsI protein, E coli; C506766:RpsL protein, E coli; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D006921:Hygromycin B; C026273:hygromycin A