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Mutagenesis of Bacteriophage T7 in vitro by Incorporation of O6-methylguanine during DNA Synthesis
L. A. Dodson, R. S. Foote, S. Mitra and W. E. Masker
Proceedings of the National Academy of Sciences of the United States of America
Vol. 79, No. 23, [Part 1: Biological Sciences] (Dec. 1, 1982), pp. 7440-7444
Published by: National Academy of Sciences
Stable URL: http://www.jstor.org/stable/13103
Page Count: 5
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An in vitro system in which bacteriophage T7 DNA is replicated and efficiently packaged into procapsids to form viable phage has been used to examine mutagenesis. The fidelity of replication was assayed both by measuring reversion of an amber mutation in an essential gene and by generation of temperature-sensitive mutants among the phage produced in vitro. Under standard reaction conditions, the fidelity of DNA replication is about equal to that normally found in vivo. However, when O6-methyldeoxyguanosine triphosphate is included in the reaction, O6-methylguanine is incorporated into newly synthesized DNA and the mutation frequencies increase 10- to 70-fold over the control. These experiments demonstrate in vitro mutagenesis with the T7 DNA replication-packaging system and provide more direct evidence for the premutagenic role of O6-methylguanine.
Proceedings of the National Academy of Sciences of the United States of America © 1982 National Academy of Sciences