188
S.M. Coelho et al.
Over the last decade, the development of molecular and, later, genomic techniques to investigate the composition of plankton communities and to study
individual phytoplankton species has had an important impact in this domain.
These techniques have provided an alternative means to study planktonic organisms, many of which cannot currently be isolated in culture. Sequencing of rDNA
cloned directly from environmental samples has proved to be a powerful tool
to study the phylogenetic diversity of eukaryotic planktonic species. This “environmental cloning” approach was originally pioneered for prokaryotic planktonic
organisms (Giovannoni et al. 1990) and it has been used much more extensively
for this group than for the eukaryotes. Nonetheless, the studies that have been carried out on planktonic eukaryotes have already revealed a remarkably high level of
diversity and have led to the discovery of several new eukaryotic groups, particularly from amongst the heterokonts and the alveolates (reviewed in Moreira and
López-García 2002). Moreover, these studies have only sampled part of the diversity present in these ecosystems and deeper sequencing is expected to reveal many
more novel sequences in the future. One of the current challenges is to link these
environmental sequences to identifiable organisms in order to gain some understanding of the ecological roles of these newly discovered species (Massana et al.
2002). For this it will be important both to develop improved culture techniques
and to improve in vivo detection methods based on techniques such as fluorescence in situ hybridisation (FISH). An important question, for example, for the
new heterokonts and alveolates mentioned above is whether they are autotrophs
or heterotrophs.
PCR amplification and cloning of rDNA sequences from environmental samples clearly allows access to a broad range of organisms. Other similar genomic
approaches are being developed, such as the use of oligonucleotide microarrays
to detect organisms and assess biodiversity in environmental samples (Medlin
et al. 2006, Metfies and Medlin 2008). However, these methods are not without
biases of their own. In particular, because they are based on PCR, the similarity
between a DNA sequence and the degenerate primers used will influence whether
that sequence is detected. In recent years, random, high-throughput sequencing of
cloned DNA fragments has been developed as an alternative approach to explore the
genetic complexity of a number of different ecosystems. Chapter 1 provides a broad
overview of how this approach is being used to explore marine systems. Here we
will concentrate on a specific example to illustrate how this approach can be used
to characterise planktonic ecosystems and its potential for the study of the algal
component of the phytoplankton.
6.4.3 Exploration of Planktonic Ecosystems Using
High-Throughput Sequencing
Craig Venter and colleagues (Venter et al. 2004, Rusch et al. 2007) used a highthroughput sequencing approach to analyse samples of micro-organisms filtered
(0.8 μm) from seawater that had been collected from several sites in the Sargasso
S.M. Coelho et al.
Over the last decade, the development of molecular and, later, genomic techniques to investigate the composition of plankton communities and to study
individual phytoplankton species has had an important impact in this domain.
These techniques have provided an alternative means to study planktonic organisms, many of which cannot currently be isolated in culture. Sequencing of rDNA
cloned directly from environmental samples has proved to be a powerful tool
to study the phylogenetic diversity of eukaryotic planktonic species. This “environmental cloning” approach was originally pioneered for prokaryotic planktonic
organisms (Giovannoni et al. 1990) and it has been used much more extensively
for this group than for the eukaryotes. Nonetheless, the studies that have been carried out on planktonic eukaryotes have already revealed a remarkably high level of
diversity and have led to the discovery of several new eukaryotic groups, particularly from amongst the heterokonts and the alveolates (reviewed in Moreira and
López-García 2002). Moreover, these studies have only sampled part of the diversity present in these ecosystems and deeper sequencing is expected to reveal many
more novel sequences in the future. One of the current challenges is to link these
environmental sequences to identifiable organisms in order to gain some understanding of the ecological roles of these newly discovered species (Massana et al.
2002). For this it will be important both to develop improved culture techniques
and to improve in vivo detection methods based on techniques such as fluorescence in situ hybridisation (FISH). An important question, for example, for the
new heterokonts and alveolates mentioned above is whether they are autotrophs
or heterotrophs.
PCR amplification and cloning of rDNA sequences from environmental samples clearly allows access to a broad range of organisms. Other similar genomic
approaches are being developed, such as the use of oligonucleotide microarrays
to detect organisms and assess biodiversity in environmental samples (Medlin
et al. 2006, Metfies and Medlin 2008). However, these methods are not without
biases of their own. In particular, because they are based on PCR, the similarity
between a DNA sequence and the degenerate primers used will influence whether
that sequence is detected. In recent years, random, high-throughput sequencing of
cloned DNA fragments has been developed as an alternative approach to explore the
genetic complexity of a number of different ecosystems. Chapter 1 provides a broad
overview of how this approach is being used to explore marine systems. Here we
will concentrate on a specific example to illustrate how this approach can be used
to characterise planktonic ecosystems and its potential for the study of the algal
component of the phytoplankton.
6.4.3 Exploration of Planktonic Ecosystems Using
High-Throughput Sequencing
Craig Venter and colleagues (Venter et al. 2004, Rusch et al. 2007) used a highthroughput sequencing approach to analyse samples of micro-organisms filtered
(0.8 μm) from seawater that had been collected from several sites in the Sargasso