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Unformatted text preview: Biology 325H Reading for April 23, 2008 Please read the following article by Yamamoto et al. in addition to BioPortal, Chapter 20, for Wednesday's lecture. Analyses We determined the population size of the bacteriophage by adding 30 m l of chloroform to a 1 ml subsample of each chemostat (to remove bacteria) and plating 100 m l of the purified subsample of bacteriophage with 300 m l of an overnight culture of bacteria in 3 ml of soft agar. We counted the number of plaques (spots cleared of bacteria by the bacteriophage) after 24 h. Because the bacteriophage were well mixed before plating, each plaque was presumed to result from a single viral particle. A lawn of ancestral bacteria (B ) was used to determine the total size of the bacteriophage population and a lawn of resistant bacteria (B 1 ) was used to calculate the number of host-range mutants (T7 1 ) in the bacteriophage population. We determined the population size of the bacteria by plating 100 m l of a subsample (without chloroform treatment) and counting the number of bacterial colonies present after 24 h of incubation. To determine the size of the B 1 population, we plated the bacteria with an equal volume of ancestral bacteriophage. To investigate local adaptation in the bacteriophage through time, we isolated two to three colonies of B 1 from the same day (day 9, 13 and 19) from each treatment during each run of the experiment. For each adaptation assay, overnight cultures of the colonies were grown up in 1 mg ml 2 1 glucose media. We plated replicate samples of T7 1 from each time point on a lawn of sympatric (from the same chemostat) and allopatric (from a different chemostat, but at the same productivity level) B 1 from the overnight culture. We used the ‘efficiency of plating’ (the number of plaques on each host) as a measure of bacteriophage infectivity. We calculated adaptation as the ratio of the number of plaques formed by the numerically dominant bacteriophage on the dominant sympatric host to the number of plaques formed on the allopatric host. Ratios of the number of plaques formed on the bacterial isolates from each replicate run of the experiment were averaged to give a mean ratio for each run. Only one replicate run of the experiment was used in assays of the closed/high productivity community due to contamination. There were often no plaques on allopatric hosts, therefore we coded the data by adding one to both the numerator and the denominator to allow calculation of a ratio. A value above one indicates local adaptation and a number below one indicates local maladaptation (that is, the number of plaques was higher on the allopatric host than on the sympatric host). All ratios were log- transformed before analysis....
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