title: The 16S rRNA phylogeny in the surface of upwelling water induced by the intrusion of Kuroshio Current
design: Approximately 50 ng of environmental DNA collected from the surface of upwelling water was used as the template for polymerase chain reaction (PCR) to specifically amplify the hypervariable V6-V9 region of 16S rRNA genes using the high-fidelity Phusion DNA Polymerase (New England Biolabs, Massachusetts, USA) and the primer set which is 16S-909F (5’-ACTCAAAKGAATWGACGG-3’) and 16S-1492R (5’-NTACCTTGTTACGACT-3’). Equimolar amounts of amplicons from two libraries were pooled for sequencing on a Roche 454 Genome Sequencer FLX+ Instrument (454 Life Sciences, Connecticut, USA).