Culturing the marine cyanobacterium Prochlorococcus

Prochlorococcus is the numerically dominant primary producer in open ocean ecosystems. Analysis of Prochlorococcus genome sequences from cultured isolates and ocean samples has broadened interest in studying this tiny cell, and efforts are underway to develop it into a model system for studying marine microbial ecology. A critical component of these efforts has been the development of systematic culturing methods that will facilitate the distribution of Prochlorococcus to diverse labs that may be interested in studying it. This paper provides detailed methods for maintaining cultures of Prochlorococcus, including a comparison of growth rates of cells on two artificial seawater media and on a standard medium that uses a natural seawater base. Procedures for agar plating, cryopreservation, obtaining new isolates, and issues associated with culture volume and carbon limitation also are described. *corresponding author; e-mail: [email protected] Authors Lisa R. Moore and Allison Coe contributed equally to this paper Acknowledgments The data collected for this manuscript is a combination of work done by various researchers, including undergraduates from the Chisholm Lab at MIT and the Moore Lab at USM, since 1988 when Prochlorococcus was first brought into culture by Brian Palenik. Although we cannot name each person who contributed, we would like to acknowledge a few who carried out some of the culture experiments: Stina Drakare and Andrew Tolonen from MIT, and Tammy Swett, Daniela Robinson, and Kristen Krumhardt from USM. This work has been supported in part by NSF (to SWC, LRM, KK, DR, MAS, TS, and EZ), DOE, the Seaver Foundation, and the Gordon and Betty Moore Foundation (to SWC). C-MORE contribution number 39. Limnol. Oceanogr.: Methods 5, 2007, 353–362 © 2007, by the American Society of Limnology and Oceanography, Inc. LIMNOLOGY and OCEANOGRAPHY: METHODS Moore et al. Prochlorococcus culturing 354 Table 1. Nutrient additions used in different Prochlorococcus growth media. AMP1 and PCR-Tu2 use artificial seawater as the base, as indicated (Turk’s Island Salt Mix). All others use filtered natural seawater as the base. Final concentrations in each medium in units of μM, unless otherwise noted. Final Concentrations CPTCK/10 PCR– PCR– Components baseda (–Cu)b PRO2c PRO99 AMP1 Tu2 d S11d