title: 454 sequencing of freshwater lake bacterioplankton: Croche Jul-08: Sample 454Reads_198
design: High-throughput sequencing of the V1ÐV3 region of the 16S rRNA gene was performed using a Roche 454 instrument with titanium chemistry by the Research and Testing Laboratory (www.researchandtesting.com, Lubbock, TX, USA). Sequencing was done with primers 28F (GAGTTTGATCNTGGCTCAG) and 519R (GTNTTACNGCGGCKGCTG).  Primers contained previously described barcodes to bio- informatically separate samples after sequencing (Hamady et al. 2008).  The PCR conditions were 95 °C for 5 min, 30 cycles of 95 °C for 30 s; 57 °C for 30 s; 72 °C for 1 min; and finally 72 °C for 7 min. The polymerase used was Roche Taq polymerase.