Approximately 340 independent wells were infected with an average of two pfu each and incubated overnight (2). Lysates were plated onto the selective strain E. coli gro89, a defective mutant of the rep gene, which encodes a DNA helicase required for particle maturation, to score for phages with the ability to infect this strain. From 12 wells, it was determined that f = 1.7 ´ 10^-5. The average final number of pfu per well was 6.6 ´ 10⁷, and B = 180 as estimated in another study (1). Thus, using Equation 1, c = log (6.6 ´ 10⁷ / 2) / log 180 = 3.3. Sequencing of 156 independent mutants showed that T = T_s = 12. Since T_s was determined, there is no selection bias due to lethals. Some bias could exist due to non-lethal effect. However, since c was small, this should not produce a large deviation in the estimate (probably less than twofold, not shown). Neglecting this effect, m_s/n/c = 3 ´ 1.7 ´ 10^-5 / 3.3 / 12 = 1.3 ´ 10^-6.

1. De Paepe, M. and F. Taddei. 2006. Viruses' life history: towards a mechanistic basis of a trade-off between survival and reproduction among phages. PLoS Biol. 4:e193.

2. Raney, J. L., R. R. Delongchamp, and C. R. Valentine. 2004. Spontaneous mutant frequency and mutation spectrum for gene A of phiX174 grown in E. coli. Environ. Mol. Mutagen. 44:119-127.