SPATIAL AND TEMPORAL SAMPLING SCALES 



For the purposes of testing the approach, spatial and temporal scales that are likely to 

 encompass the greatest change in the activity of Synechococcus populations would be most 

 valuable. Thus, a seasonal series of short cruises (e.g. spring, summer, fall) would probably be 

 preferred. Within each cruise, cross-shelf depth profile transects would be appropriate. In 

 addition, 24- or 48-hour "diel" stations during which the water column could be sampled 

 intensively (e.g. every 2-3 hours) would be invaluable for investigating the changes in population 

 characteristics that are driven by the light:dark cycle. 



METHODS AND PLATFORMS 



Method development is a major part of this project, as reviewed briefly above. Our 

 anticipated strategy for cruises would involve preservation of samples at sea, and detailed analysis 

 of these samples back on shore. Therefore, we do not expect to require any specialized 

 platforms. Nevertheless, for the purpose of developing sampling strategies during the cruise, and 

 for routine monitoring of incubation experiments, we may request space for a ship-board flow 

 cytometer. 



STRENGTHS AND LIMITATIONS OF PROPOSED RESEARCH 



The strengths of the approach we propose derive both from the independence of the 

 method from the need for any incubation, and from the analytical power of flow cytometry. 

 Although incubations are a necessary part of many biological oceanographic measurements, and 

 the data derived from these measurements forms the basis for much of our knowledge about the 

 regulation of microbial and phytoplankton communities, the potential artifacts that such 

 incubations introduce are well known and widely recognized. By using the "biochemical state" 

 of the organisms recovered from the natural environment as an estimator for growth rate, our 

 approach holds out the promise of measuring in situ growth rates that are completely without 

 incubation artifacts. The application of flow cytometry to this problem not only provides for 

 rapid, quantitative, and objective analysis, but also allows such analysis to be made on a 

 group-specific basis. Flow cytometry furthermore can be used to physically sort cells out of the 

 sample, allowing further microscopic, biochemical or molecular analysis of the organisms of 

 interest 



The major weakness of the approach is its dependance upon a uniformity in the 

 relationship between the biochemical index of choice and growth rate under different 

 environmental conditions, and among all the organisms of interest. Our laboratory experiments 

 are designed to ascertain the degree to which this assumption is met among the Synechococcus. 

 If the growth rate response is modulated by specific environmental parameters (e.g. temperature), 



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