Genotypic reversion by methylene Blue: The orientation of guanine-hydroxymethylcytosine at mutated sites in rII mutants of phage T4

Summary Methylene Blue (MB) induced reversion of rII mutants to which a GC base pair was previously assigned is investigated, both in K₁₂() and in B bacteria. The rII mutants used are the same ones whose hydroxylamine (HA) induced reversion rates have previously been investigated by Levisohn (1967) in a similar series of experiments.The results show a relationship between MB-induced and HA-induced reversion patterns which can be interpreted on the assumptions: 1. Both MB and HA produce transitions, but MB acts on Guanine (in agreement with Simon and van Vunakis 1962), while HA acts on Cytocine; 2. Only one injected DNA strand (genetic strand) transmits hereditary information to the progeny while the other strand (associated strand) is disintegrated soon after performing some early functions and its nucleotides are re-used in the formation of new DNA; 3. When the injected associated strand disintegrates, Cytocine bases altered by HA can be re-used, while Guanine bases altered by MB cannot.The assumption that only one strand of the injected DNA transmits hereditary information to the progeny (Simplex hypotheses) is also used in a new interpretation of the mutagenic-induced reversion pattern of lysozyme mutants. This new interpretation avoids a difficulty in the earlier interpretation proposed by Terzaghi, Streisinger and Stahl (1962) which assigned an AT base pair to a mutant strongly HA revertible, while none of the mutants to which a GC base pair had been assigned was HA revertible.The main advantage of the Simplex interpretations of mutagenic phenomena is that self-consistent base assignments to each mutant can be made following a series of rules which have no exceptions: 1. An early function mutant with a high HA reversion rate in non-permissive host can consistently be assigned an odd C in the genetic strand (if odd designates a base which, when replaced through a transition, causes reversion to wild type); 2. A mutant with high MB reversion rate can consistently be assigned an odd G in the genetic strand; 3. A few mutants which present both of these properties are assigned both an odd C and an odd G in a genetic strand triplet; 4. A high HA reversion rate in permissive host but not in non-permissive host is consistently indicative of an odd G with no odd C in the genetic strand triplet.This work was supported by research grant GM-12581 from the Division of General Medical Sciences of the National Institutes of Health, United States Public Health Service.