Pyrosequencing of nitrous oxide reductase gene nosZ: agricultural soil
DNA was extracted from soil using the Fast DNA Spin Kit for Soil (MP Biomedicals, Santa Ana, USA). A two-step PCR was performed to amplify the nosZII (nosZIIF/nosZIIR; Jones et al., ) genes to introduce the barcode, sequencing key and adapter. Pyrosequencing was performed by Microsynth (Balgach, Switzerland) on a Roche’s 454 FLX Genome Sequencer using Titanium chemistry. Sequence data was initially screened and de-multiplexed using QIIME (version 1.8.0, Caporaso et al., 2010) with the default parameters.