Metagenomic sequences from DNA from pico-planktonic 5-0.2 micrometer size fraction of the deep chlorophyll maximum
No library was constructed for the direct sequencing approach where the environmental DNA was sent directly for sequencing by 454 ( single, unpaired reads ).  The library construction protocol for the the preparation of the fosmid library from the environmental DNA is described here briefly. The fosmid genomic library was constructed using the CopyControl™ Fosmid Library Production Kit (Epicentre), as described by the manufacturer's instructions, from approximately 9.7 micrograms of DNA. The fosmid library generated had a total of 12192 clones. From this collection 1152 (twelve 96 well plates) were randomly selected for 454 pyrosequencing. Clones were individually grown and induced using 1ml 96 well-plates. All wells within each plate we pooled before DNA extraction.  DNA was extracted using the QIAprep Spin Miniprep kit (QIAGEN). 5 microgram of DNA from each pooled plate was sent separately for sequencing (Roche 454 GS-FLX system, Titanium chemistry, by GATC, Konstanz, Germany). DNA from each plate was tagged individually using a multiplex identifier adaptor (MID) containing a unique 10-base pair sequence that is recognized by the sequencing analysis software, allowing for automated sorting of MID-containing reads and facilitating the assembly.