Induction of mutation in vitro in phage phi X174 am3 by N4-aminodeoxycytidine triphosphate.
When phi X174 am3-phage-infected E. coli is treated with N4-aminocytidine, reversion of the phage to the wild type is efficiently induced. The mechanism of this reversion is considered to consist of metabolic conversion of N4-aminocytidine into its deoxynucleoside 5'-triphosphate followed by incorporation of the nucleotide into the replicating phage DNA, thereby causing AT-to-GC transition at the am3 locus. The second half of this mechanism has now been experimentally proved, using an in vitro mutagenesis system. Thus, by nick-translation, N4-aminodeoxycytidine 5'-triphosphate was incorporated into the replicative form of phi X174 am3 DNA, and the DNA was used to transfect CA++-treated E. coli HF4714 (sup+). The reversion frequency of the phage produced was up to one-order of magnitude greater than that of the control in which the nick-translation had been done without the addition of N4-aminodeoxycytidine triphosphate. This nucleotide analog may be useful as a reagent for in vitro site-directed mutagenesis.
Takahashi
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M
Negishi
K
K
Hayatsu
H
H
D010584:Bacteriophage phi X 174 / Q000235:genetics*; D001482:Base Composition; D001711:Biotransformation; D004279:DNA, Viral / Q000235:genetics; D003845:Deoxycytosine Nucleotides / Q000378:metabolism / Q000633:toxicity*; D004926:Escherichia coli / Q000235:genetics; D009152:Mutagenicity Tests; D009153:Mutagens; D014176:Protein Biosynthesis; D014162:Transfection
D004279:DNA, Viral; D003845:Deoxycytosine Nucleotides; D009153:Mutagens; C046787:N(4)-amino-2'-deoxycytidine triphosphate