Preliminary experiments suggest that these cells show contact inhibition of 
replication in colonies containing at least 100 cells, as well as in dense 
monolayers (35). All such clones tested to date appear stable in their ability to 
express the apparent contact-inhibited state. 
Mutation Frequency and Expression Time 
Mutation is a complex biological process involving much more than 
interaction of mutagen with DNA, or more generally, with the DNA-replicating 
system. The mutagen-DNA interaction usually results not in mutation, but in 
the creation of premutational lesions which become expressed as mutations 
only after a series of additional events has occured (6). For example, 
auxotrophic mutants usaully become expressed as soon as the cells become 
depleted of normal gene product. The length of time required for this and 
other preliminary events to occur may vary with the nature of the gene 
product, the specific mutagen utilized, and the doses employed (5, 7). To 
determine the influence of expression time on observed mutation frequency 
for the mutant phenotypes reported above, experiments employing variable 
expression time were carried out with EMS and BrdU. Doses used were those 
giving maximal observed mutation frequency when the expression time was 
Five days (3 x 10"^ molar for EMS, 1 x lcH - for BrdU). Expression time was 
varied between two and eleven days. The data are given in Table 74, again in 
terms of the parameters of equation [2]. Observed mutation frequency is 
plotted in Figure 7-8 as a function of expression time. 
Optimal expression time is defined as the interval between mutagen 
quenching, and the application of selective conditions yielding maximal 
mutation frequency when dose is held constant. This is observed to be five 
days for BrdU and eight days for EMS. Moreover, for a two-day expression 
period, statistically significant numbers of mutants could always be identified 
in populations exposed to EMS, whereas none are found in populations treated 
with BrdU. Once optimal expression time is exceeded, mutant cell frequency 
decreased rapidly, suggesting that these types of mutants are at a replicative 
disadvantage relative to wild-type cells under nonselective conditions. The fact 
that optimal expression time was different for the two mutagens when tested 
at doses showing similar toxicity, suggest that expression time may be mutagen 
dependent. This leads support to the contention that assays utilizing variable 
expression time shall be required when screening compounds for mutagenic 
activity (4). This may be particularly important for weak mutagens. 
DISCUSSION 
The genetic toxicology of inorganic compounds has largely been ignored, 
despite the fact that many are ubiquitous, highly toxic, and implicated as 
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