This sponge EMP blast database has been created for a recent publication. If you use the database, please cite this article:
Dat TTH, Steinert G, Thi Kim Cuc N, Smidt H, Sipkema D. (2018) Archaeal and bacterial diversity and community composition from 18 phylogenetically divergent sponge species in Vietnam. PeerJ 6:e4970 https://doi.org/10.7717/peerj.4970
SpongeEMP deblurred subOTUs BLAST database
The sponge microbiome project (see https://academic.oup.com/gigascience/article/6/10/1/4082886) deblurred biom file "final.withtax.biom" was downloaded from https://github.com/amnona/SpongeEMP/tree/master/sponge_emp/data
The biom file was converted to a tsv file:
biom convert -i final.withtax.biom -o final.withtax.tsv --to-tsv --header-key taxonomy
Subsequently, the stored OTU representative sequence information was extracted from this text file and stored in a separated fasta file "final.withtax.v12.fa"
Representative sequences for each deblurred OTU was classified using mothur v.1.39.5 https://www.mothur.org/wiki/Download_mothur and silva128 https://www.mothur.org/wiki/Silva_reference_files#Release_128
a) First just a simple classification without chimera search and removal of Mitochondria-Chloroplast-Eukaryota-unknown sequences
classify.seqs(fasta=final.withtax.v12.fa, template=silva.nr_v128.align, taxonomy=silva.nr_v128.tax, cutoff=80, probs=T, processors=6)
taxlevel rankID taxon daughterlevels total 0 0 Root 4 83908 1 0.1 Archaea 10 1025 1 0.2 Bacteria 53 64429 1 0.3 Eukaryota 13 915 1 0.4 unknown 1 17539
b) Chimera search Creating name file to use EMP sequences as template for chimera search
unique.seqs(fasta=final.withtax.v12.fa)
Chimera search and removal
chimera.vsearch(fasta=final.withtax.v12.fa, name=final.withtax.v12.names, dereplicate=t, processors=6)
Using 6 processors. Checking sequences from final.withtax.v12.fa ... When using template=self, mothur can only use 1 processor, continuing. vsearch v2.3.4_linux_x86_64, 15.6GB RAM, 8 cores https://github.com/torognes/vsearch
Reading file final.withtax.v12.temp 100%
8390800 nt in 83908 seqs, min 100, max 100, avg 100
Masking 100%
Sorting by abundance 100%
Counting unique k-mers 100%
Detecting chimeras 100%
Found 0 (0.0%) chimeras, 83908 (100.0%) non-chimeras,
and 0 (0.0%) borderline sequences in 83908 unique sequences.
Taking abundance information into account, this corresponds to
0 (0.0%) chimeras, 83908 (100.0%) non-chimeras,
and 0 (0.0%) borderline sequences in 83908 total sequences.
It took 27 secs to check 83908 sequences. 0 chimeras were found.
c) emoval of certain sequences Removing Mitochondria-Chloroplast-Eukaryota-unknown sequences from seq file
remove.lineage(fasta=final.withtax.v12.fa, taxonomy=final.withtax.v12.nr_v128.wang.taxonomy, taxon=Mitochondria-Chloroplast-Eukaryota-unknown)
Output File Names: final.withtax.v12.nr_v128.wang.pick.taxonomy final.withtax.v12.pick.fa
final.withtax.v12.fa 83908 OTUs final.withtax.v12.pick.fa 64424 OTUS
a) Go to the BLAST download folder and download the BLAST executables for your operating system: ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST/ - unzip this archive.
b) Copy the fasta file into the ncbi-blast-x.x.x bin folder.
c) Open your terminal and go to the bin folder. Use the following command to create the spongeemp nucleotide BLAST database:
makeblastdb -in final.withtax.v12.pick.fa -out DeblurV12PickOTUsSilva128 -dbtype 'nucl' -hash_index