Amplicon sequencing of prokaryotic 16S rRNA gene and nosZ in agricultural soil

DNA and RNA molecules were extracted from Japanese agricultural soils, and the RNA was transcribed to cDNA by reverse transcription reaction using random 6-mer primers. The extracted DNA was subjected to PCR amplification of prokaryotic 16S rRNA gene using the oligonucleotide primers 515F and 806R, whereas the synthesized cDNA was used for PCR amplification of the gene encoding nitrous oxide reductase (NosZ) using the primer nosZ-123-145F and nosZ-481-499-R primers. The PCR amplicons were purified and subjected to the sequencing using the illumina MiSeq sequencer and V2 reagent kit.