title: Microbial community (bacteria 16S rRNA V4V5 region) of forest soil and biochar amended soil in Pacific Northwest USA
design: Sequencing and bioinformatics work was done by the IBEST staff at the University of Idaho IBEST facility (Moscow, ID).  DNA amplicons were sequenced on a paired-end 300bp Illumina MiSeq platform (Illumina, Inc., San Diego, CA) in a four file format: Read 1, Index Read 1, Index Read 2, and Read 2.  The custom python application dbcAmplicons (https://github.com/msettles/dbcAmplicons) was used to assign reads.  Raw fastq files were screened for presence of template-specific primer sequences (V1V3, V4V5, LSU) and separated by primer sequence (maximum allowed mismatches 4) and index sequence (maximum allowed mismatches 1).  Sequences were overlapped using Flash (maximum proportion of mismatch 0.25=number of mismatches/length of overlap) and then classified with the RDP Classifier using the 16S database for V1V3 and V4V5 and the fungal LSU database for the LSU sequences.  Bootstrap cutoff was 0.5.  The resulting fixrank file for each amplicon target was parsed to generate abundance of taxa by proportion and number of reads per sample as well as summaries of taxonomic classifications for each target.