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mothur_fungal
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#Pipeline desarrollado por Estefanny Gutiérrez
# Agosto 2021
make.file(inputdir=., type=fastq, prefix=stability)
make.contigs(file=stability.files, processors=8)
summary.seqs(fasta=stability.trim.contigs.fasta)
screen.seqs(fasta=stability.trim.contigs.fasta, group=stability.contigs.groups, summary=stability.trim.contigs.summary, minlength=27, maxlength=275)
unique.seqs(fasta=stability.trim.contigs.good.fasta)
count.seqs(name=stability.trim.contigs.good.names, group=stability.contigs.good.groups)
summary.seqs(count=stability.trim.contigs.good.count_table)
pre.cluster(fasta=stability.trim.contigs.good.unique.fasta, count=stability.trim.contigs.good.count_table, diffs=2)
chimera.vsearch(fasta=stability.trim.contigs.good.unique.precluster.fasta, count=stability.trim.contigs.good.unique.precluster.count_table, dereplicate=t)
remove.seqs(fasta=stability.trim.contigs.good.unique.precluster.fasta, accnos=stability.trim.contigs.good.unique.precluster.denovo.vsearch.accnos)
summary.seqs(fasta=stability.trim.contigs.good.unique.precluster.pick.fasta, count=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.count_table)
# En lugar de trainset se usa la Database de UNITE (IMPORTANTE: Se usa en FASTA Y TAX de no hacerlo marcará error de database no alineada!)
classify.seqs(fasta=stability.trim.contigs.good.unique.precluster.pick.fasta, count=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.count_table, reference=database.fasta, taxonomy=database.tax, cutoff=80)
# Database se cambia por la base de datos UNITE
remove.lineage(fasta=stability.trim.contigs.good.unique.precluster.pick.fasta, count=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.count_table, taxonomy=stability.trim.contigs.good.unique.precluster.pick.database.wang.taxonomy, taxon=Chloroplast-Mitochondria-unknown-Archaea-Bacteria)
# Alternativamente se puede hacer esto:
# Database se cambia por la base de datos UNITE
remove.lineage(fasta=stability.trim.contigs.good.unique.precluster.pick.fasta, count=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.count_table, taxonomy=stability.trim.contigs.good.unique.precluster.pick.database.wang.taxonomy, taxon=Chloroplast-Mitochondria-Archaea-Bacteria)
summary.tax(fasta=current, count=current)
make.shared(count=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.count_table)
# Database se cambia por la base de datos UNITE
classify.otu(list=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.ASV.list, count=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.count_table, taxonomy=stability.trim.contigs.good.unique.precluster.pick.database.wang.pick.taxonomy, label=asv)
make.biom(shared=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.ASV.shared, constaxonomy=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.ASV.ASV.cons.taxonomy)
rarefaction.single(shared=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.ASV.shared, calc=sobs, freq=100)
summary.single(shared=stability.trim.contigs.good.unique.precluster.denovo.vsearch.pick.pick.ASV.shared, calc=nseqs-coverage-sobs-invsimpson-shannon-chao, subsample=T)