Jochem, F. J.. Institut fuer Meereskunde, Frank.Jochem@mare.de
Smith, G. J.. Hopkins Marine Station, symbios@aol.com
Gao, A. RUTGERS University of New Jersey, gao@aesop.rutgers.edu
Robertson, D. Harvard University, deb@biosun.harvard.edu
Cabello-Pasini, A. Instituto de Investigaciones Oceanologicas, acabello@faro.ens.uabc.mx
Kohrs, D. G.. Moss Landing Marine Laboratory,
Alberte, R. S.. Arete Associates, alberte@arete-dc.com

 
IN-SITU PROBING FOR PHYTOPLANKTON GROWTH AND ENZYME ACTIVITY IN SINGLE CELLS - FLOW CYTOMETRY OF DNA CELL CYCLES AND NITRATE REDUCTASE
 
Intrinsic growth rates are essential to understand phytoplankton dynamics and physiological responses upon environmental changes. Cell cycle analysis provides growth rates without biasing incubations. The blue-light excitable DNA dye YoYo-1 suitable for application on benchtop cytometers accommodated high resolution DNA analysis while preserving chlorophyll autofluorescence for discrimination of algal cells against other particles. Diel patterns of DNA replication and cell division were mirrored by chlorophyll signals and could be related to the light-cycle in cultures of various algal species and changes in nitrogen supply in the marine diatom Skeletonema costatum. Nitrate reductase (NR), a critical enzyme in the nitrate metabolism, is an extremely low abundant and transiently expressed protein. Even during periods of peak nitrate assimilation the NR protein (NRP) is present at less than 10 fg/cell. Using affinity-purified polyclonal antibodies raised against NRP from S. costatum, changes in NRP abundance were monitored following a transition from low to high f-ratio growth of S. costatum in a chemostat system. Accumulation of NRP could be clearly visualized by fluorescence microscopy or Western Blots 48 hrs after the transition. Cytometric analysis of antiNR-GAR-FITC fluorescence, however, enabled detection of short term (<12 hrs) changes in NRP in response to the increased nitrate supply as well as low NRP levels during low f-ratio growth.
 
Day: Tuesday, Feb. 2
Time: 03:30 - 03:45pm
Location: Sweeney Center
 
Code: SS41TU0330S