Targeted mutagenesis of DNA with alkylating RecA assisted oligonucleotides.
Site-specific mutation was demonstrated in a shuttle vector system using nitrogen mustard-conjugated oligodeoxyribonucleotides (ODNs). Plasmid DNA was modified in vitro by ODNs containing all four DNA bases in the presence of Escherichia coli RecA protein. Up to 50% of plasmid molecules were alkylated in the targeted region of the supF gene and mutations resulted upon replication in mammalian cells. ODNs conjugated with either two chlorambucil moieties or a novel tetrafunctional mustard caused interstrand crosslinks in the target DNA and were more mutagenic than ODNs that caused only monoadducts.
Singer
M J
MJ
Epoch Pharmaceuticals, Inc., 12277 134th Ct. NE, Redmond, WA 98052, USA. msinger@epochpharm.com
Podyminogin
M A
MA
Metcalf
M A
MA
Reed
M W
MW
Brown
D A
DA
Gamper
H B
HB
Meyer
R B
RB
Wydro
R M
RM
D000477:Alkylating Agents / Q000378:metabolism*; D000478:Alkylation; D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D016153:Genes, Suppressor; D005822:Genetic Vectors; D008466:Mechlorethamine / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D010957:Plasmids; D012343:RNA, Transfer / Q000235:genetics; D011938:Rec A Recombinases / Q000378:metabolism*; D017422:Sequence Analysis, DNA
D000477:Alkylating Agents; D009838:Oligodeoxyribonucleotides; C096275:supF tRNA; D008466:Mechlorethamine; D004247:DNA; D012343:RNA, Transfer; D011938:Rec A Recombinases