Increased SFHR gene correction efficiency with sense single-stranded DNA.
BACKGROUND
The correction of a mutated gene by the small fragment homologous replacement (SFHR) method is a highly attractive approach for gene therapy. However, the current SFHR method with a heat-denatured double-stranded PCR fragment yielded a low correction efficiency.


METHODS
Single-stranded (ss) DNA fragments were prepared from ss phagemid DNA and tested in a gene correction assay with an inactivated Hyg-EGFP fusion gene, as a model target.


RESULTS
A 606-nt sense, ss DNA fragment dramatically (12-fold) improved the gene correction efficiency, although the antisense strand showed only minimal correction efficiency.


CONCLUSIONS
These results suggest that the use of a sense, single-stranded DNA fragment is useful in the SFHR method for the correction of mutated genes.
Tsuchiya
Hiroyuki
H
Laboratory for Molecular Design of Pharmaceutics and COE Program in the 21st Century, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita-12, Nishi-6, Sapporo, Hokkaido 060-0812, Japan.
Harashima
Hideyoshi
H
Kamiya
Hiroyuki
H
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D017931:DNA Primers; D004277:DNA, Single-Stranded / Q000235:genetics*; D010957:Plasmids; D016133:Polymerase Chain Reaction
D017931:DNA Primers; D004277:DNA, Single-Stranded