Representative model organisms form the basis on which biology is constructed, and
pure cultures offer many opportunities for discovery. However, our view of the importance of
axenic cultures changed dramatically at the turn of the last century upon realizing that the majority
of environmentally relevant microbes still remains uncultured. The sequencing revolution has
led us to a point where we can identify the microbial world in which we live, but many questions
remain regarding the autecology of planktonic microbes and their interactions with their environment.
Thus, it is essential to isolate and cultivate the key microbial players to gain a deeper insight
into their ecology. If the past is a guide, the way forward in confronting the so-called ‘great plate
count anomaly’ is the use of more subtle and refined approaches to culturing, using a number of
methods and processes that are now becoming available. The vast amount of information accumulated
from genome sequencing alone has yet to result in the isolation of the most important and
abundant microbes of aquatic systems. We highlight the merits of pure cultures and discuss the
critical need to integrate information from a variety of different sources to isolate planktonic
microbes. We also describe how to culture bacteria of interest with a full cycle isolation approach
based on targeted enrichment and illustrate the benefits of pure cultures with 2 examples of
isolated representatives of freshwater Betaproteobacteria.