Mutagenesis by aflatoxin in M13 DNA: Base-substitution mechanisms and the origin of strand bias

Sudhir Sahasrabudhe, Kumar Sambamurti, M. Zafri Humayun

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The two goals of the experiments described here are: (a) to examine whether there is a strand bias in mutagenic processing of bulky lesions in M13 replicative form (RF) DNA, and (b) to examine the mutational mechanisms of metabolically activated aflatoxin. For these experiments, two types of nicked heteroduplex M13 RF DNA molecules (+WT/-am1 and +am1/-WT) in which either the minus (-) or the plus (+) strand carried a gene 1 amber nonsense codon, were constructed. Heteroduplex DNAs were modified in vitro with aflatoxin B1 activated by hamster liver S9 enzymes, and transfected into SOS(UV)-induced Escherichia coli (Supo/uvrA-/mucAB+). Forward mutations in the lacZ α-complementing gene segment were scored and sequenced. Results indicated that aflatoxin-induced mutation frequencies in the +WT/-am1 heteroduplex were significantly greater than those in the +am1/-WT heteroduplex, suggesting more efficient mutagenic processing of lesions in the plus strand. These results permit specific suggestions for improved mutation detection in the extensively used M13 forward mutagenesis system. Sequence analysis of point mutations from the +WT/-am1 experiments showed that most substitutions were targeted to plus-strand guanines. Both G-to-A transitions and G-to-T transversions were induced with equal effeciency. Since activated aflatoxin B1 is known to react almost exclusively with DNA guanines at the N7 position, these results suggest that bulky lesions at guanine N7 position may have the properties of mis-instructional as well as non-instructional lesions.

Original languageEnglish (US)
Pages (from-to)20-25
Number of pages6
JournalMgg Molecular & General Genetics
Issue number1
StatePublished - May 1989

All Science Journal Classification (ASJC) codes

  • Genetics


  • Aflatoxin
  • Bulky DNA adducts
  • DNA replication
  • Lesion bypass
  • Mutagenic mechanisms


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