Targeted gene conversion in a mammalian CD34+-enriched cell population using a chimeric RNA/DNA oligonucleotide.
Gene conversion of genetically inherited point mutations is a fundamental methodology for treating a variety of diseases. We tested the feasibility of a new approach using an RNA/DNA chimeric oligonucleotide. The beta-globin gene was targeted at the point mutation causing sickle cell anemia. The chimera is designed to convert an A residue to a T after creating a mismatched basepair. In a CD34+-enriched population of normal cells a 5-11% conversion rate was measured using restriction enzyme polymorphism and direct DNA sequence analyses. The closely related delta-globin gene sequence appeared unchanged despite successful conversion at the beta-globin locus.
Xiang
Y
Y
Department of Pharmacology, Kimmel Cancer Institute, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Cole-Strauss
A
A
Yoon
K
K
Gryn
J
J
Kmiec
E B
EB
D018952:Antigens, CD34 / Q000235:genetics*; D004247:DNA / Q000235:genetics*; D005785:Gene Conversion; D018390:Gene Targeting / Q000379:methods; D005822:Genetic Vectors / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed / Q000235:genetics*; D016376:Oligonucleotides, Antisense / Q000235:genetics*; D012313:RNA / Q000235:genetics*; D017422:Sequence Analysis, DNA
D018952:Antigens, CD34; D016376:Oligonucleotides, Antisense; D012313:RNA; D004247:DNA