Hey guys,
I sequenced some environmental samples using miseq and I ran the data through a QIIME pipeline. After demultiplexing, OTU picking, chimera removal and taxonomic assignment using pair-end reads, I have a lot of NA samples (up to 50%). I was advised to recover nonassigned OTUs and blast the fasta file generated, then include these into the taxonomy. However, I'm unsure what I should do with the .txt file generated from blasting the NA OTUs.
As an aside, I tried using just the reverse reads and the amount of NA OTUs was extremely low (~2%) but I'm not sure this is an accurate representation of what's really in my samples.
If anyone has some suggestions I'd be grateful. I included the commands I entered below.
Thanks!
OTU_picking_pair_end_read -m mapping_file_nbg_70bp.txt -l nbg -a 0.1 -c 0.900 -f /home/javier/16S_analysis/db/primer_27F_70_nt_db -r /home/javier/16S_analysis/db/primer_338R_70_nt_db -o 70 -e 70 -t /home/javier/16S_analysis/db/Silva_111_taxa_map_RDP_6_levels_full.txt -q 2 -k 100
OTU_recovery_for_reassignment -t tax_files -n NA_otus_nbg_70bp -u NA_unique_otus_nbg_70bp -a alternative_taxonomy_NA_otus_nbg_70bp -p NA
taxonomical_reassignment -a alternative_taxonomy_NA_otus_nbg_70bp1 -t tax_files
