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main.nf
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#!/usr/bin/env nextflow
/*
* Copyright (c) 2017-2022, Centre for Genomic Regulation (CRG)
*
* Copyright (c) 2017, Anna Vlasova
*
* Copyright (c) 2017, Emilio Palumbo
*
* Copyright (c) 2018-2022, Toni Hermoso Pulido
*
* Functional Annotation Pipeline for protein annotation from non-model organisms
* from Genome Annotation Team in Catalonia (GATC) implemented in Nextflow
*
* This program is free software: you can redistribute it and/or modify
* it under the terms of the GNU General Public License as published by
* the Free Software Foundation, either version 3 of the License, or
* (at your option) any later version.
*
* This program is distributed in the hope that it will be useful,
* but WITHOUT ANY WARRANTY; without even the implied warranty of
* MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the
* GNU General Public License for more details.
*
* You should have received a copy of the GNU General Public License
* along with this program. If not, see <http://www.gnu.org/licenses/>.
*/
// there can be three log levels: off (no messages), info (some main messages), debug (all messages + sql queries)
// default parameters
params.help = false
params.debug = false
params.dbEngine = "SQLite" // MySQL otherwise
params.mysqllog = "${baseDir}/tmp"
// Main input files
params.proteinFile = null;
params.gffFile = null;
params.speciesName = "organism"
params.dbname = "organismDB"
// Main result and log dirs
params.resultPath = "${baseDir}/results/"
params.stdoutLog = "${baseDir}/logs/out.log"
params.stderrLog = "${baseDir}/logs/err.log"
// Sizes for different programs
params.chunkIPSSize = null
params.chunkBlastSize = null
params.chunkKoalaSize = null
params.chunkWebSize = null
params.debugSize = 2
// Blast related params
params.blastFile = null
params.evalue = 0.00001
params.diamond = null
params.blastDbPath = null
// GO retrieval params
params.gogourl = ""
params.gogohits = 30
params.blastAnnotMode = "common" // common, most, all available so far
// KEGG
params.kolist = null
params.koprofiles = null
params.koentries = null
params.kegg_release = null
params.kegg_species = "hsa, dme, cel, ath"
params.keggFile = null
// Params for InterProScan
// Temporary location for InterproScan intermediary files. This can be huge
params.ipscantmp = "${baseDir}/tmp/"
// Location of InterproScan properties. Do not modify unless it matches your container
params.ipscanproperties = "/usr/local/interproscan/interproscan.properties"
params.ipscandata = ""
ipscandata = "/usr/local/interproscan/data"
if ( params.ipscandata != "" ) {
ipscandata = params.ipscandata
} else {
if ( params.iprscanVersion ) {
if ( params.dbPath ) {
if ( new File( params.dbPath + "/iprscan/" + params.iprscanVersion ).exists() ) {
ipscandata = "${params.dbPath}/iprscan/${params.iprscanVersion}"
}
}
}
}
// Params for dealing with GFF
params.gffclean = true
params.gffstats = true
// Remove version from protein entries (e.g. X5543AP.2)
params.rmversion = false
// File with GO information, otherwise is downloaded
params.oboFile = null
// Skip params
params.skip_cdSearch = false
params.skip_sigtarp = false
// Mail for sending reports
params.email = ""
// Download paths
params.dbPath = null
params.blastDbFolder = null
params.dbipscanPath = null
params.dbKOPath = null
// Handling defaults from download part
Date date = new Date()
String datePart = date.format("yyyyMM")
// Handle BlastDB paths
blastDbPath = null;
if ( ! params.blastDbPath ) {
if ( ! params.blastDbFolder ) {
if ( ! params.dbPath ) {
log.info "At least dbPath needs to be defined!"
exit 1
} else {
blastDbFolder = "${params.dbPath}/ncbi/${datePart}/blastdb/db"
// Let's assume Swissprot
blastDbPath = "${blastDbFolder}/swissprot"
}
} else {
// Let's assume Swissprot
blastDbPath = "${params.blastDbFolder}/swissprot"
}
} else {
blastDbPath = params.blastDbPath
}
// KEGG paths
kolist = null
koprofiles = null
koentries = null
if ( params.kolist ) {
kolist = params.kolist
}
if ( params.koprofiles ) {
koprofiles = params.koprofiles
}
if ( params.koentries ) {
koentries = params.koentries
}
if ( ! params.kolist && ! params.keggFile ) {
if ( params.koVersion && params.dbPath ) {
kolist = "${params.dbPath}/kegg/${params.koVersion}/ko_list"
}
}
if ( ! params.koprofiles && ! params.keggFile ) {
if ( params.koVersion && params.dbPath ) {
koprofiles = "${params.dbPath}/kegg/${params.koVersion}/profiles"
}
}
if ( ! params.koentries ) {
if ( params.koVersion && params.dbPath ) {
koentries = "${params.dbPath}/kegg/${params.koVersion}/ko_store"
}
}
//print usage
if ( params.help ) {
log.info ''
log.info 'Functional annotation pipeline'
log.info '----------------------------------------------------'
log.info 'Run functional annotation for a given species.'
log.info ''
log.info 'Usage: '
log.info " ./nextflow run main.nf --config params.config [options]"
log.info ''
log.info 'Options:'
log.info '-resume resume pipeline from the previous step, i.e. in case of error'
log.info '-help this message'
exit 1
}
/*
* Parse the input parameters
*/
// species-specific parameters
protein = null
annotation = null
config_file = file(params.config)
dbFile = false
boolean exists = false
boolean mysql = false
gffavail = false
gffclean = false
gffstats = false
// Skip
skip_cdSearch = false
skip_sigtarp = false
if( params.dbEngine.toLowerCase()=="mysql" ) {
mysql = true
}
if ( params.gffclean ) {
gffclean = true
}
if ( params.gffstats ) {
gffstats = true
}
if ( params.skip_cdSearch ) {
skip_cdSearch = true
}
if ( params.skip_sigtarp ) {
skip_sigtarp = true
}
// Handling MySQL in a cleaner way
dbhost = null
// Getting contents of file
if ( mysql ) {
dbhost = "127.0.0.1" // Default value. Localhost
if ( new File( params.mysqllog+"/DBHOST" ).exists() ) {
dbhost = new File( params.mysqllog+"/DBHOST" ).text.trim()
}
} else {
dbFileName = params.resultPath+params.dbname+'.db'
dbFile = file(dbFileName)
if ( dbFile.exists() && dbFile.size() > 0 ) {
exists = true
}
}
// print log info
log.info ""
log.info "Functional annotation pipeline"
log.info ""
log.info "General parameters"
log.info "------------------"
if ( params.proteinFile == null || params.proteinFile == "" ) {
log.info "No protein sequence file specified!"
exit 1
} else {
if ( file( params.proteinFile ).exists() && file( params.proteinFile ).size() > 0 ) {
log.info "Protein sequence file : ${params.proteinFile}"
protein = file(params.proteinFile)
} else {
log.info "Protein sequence file does not exist or it is empty!"
exit 1
}
}
if ( params.gffFile == null || params.gffFile == "" ) {
log.info "No GFF Structural Annotation file specified!"
log.info "We proceed anyway..."
} else {
if ( file( params.gffFile ).exists() && file( params.gffFile ).size() > 0 ) {
log.info "GFF Structural Annotation file : ${params.gffFile}"
gffavail = true
annotation = file(params.gffFile)
} else {
log.info "GFF Structural Annotation file is missing or empty."
log.info "We stop the pipeline so you can check it and define as \"\" otherwise if no GFF file is provided."
exit 1
}
}
if ( params.blastFile ) {
log.info "BLAST results file : ${params.blastFile}"
}
log.info "Species name : ${params.speciesName}"
log.info "KEGG species : ${params.kegg_species}"
if ( mysql ) {
log.info "MySQL FA database : ${params.dbname}"
} else {
log.info "SQLite FA database : $dbFileName"
}
if ( skip_cdSearch ) {
log.info "CD-Search queries will be skipped."
}
if ( skip_sigtarp ) {
log.info "SignalP and targetP queries will be skipped."
}
// split protein fasta file into chunks and then execute annotation for each chunk
// chanels for: interpro, blast, signalP, targetP, cdsearch_hit, cdsearch_features
chunkSize = params.chunkSize
chunkBlastSize = chunkSize
chunkIPSSize = chunkSize
chunkKoalaSize = chunkSize
chunkWebSize = chunkSize
if ( params.chunkBlastSize ) {
chunkBlastSize = params.chunkBlastSize
}
if ( params.chunkIPSSize ) {
chunkIPSSize = params.chunkIPSSize
}
if ( params.chunkKoalaSize ) {
chunkKoalaSize = params.chunkKoalaSize
}
if ( params.chunkWebSize ) {
chunkWebSize = params.chunkWebSize
}
seqData = Channel
.from(protein)
.splitFasta( by: chunkSize )
seqBlastData = Channel
.from(protein)
.splitFasta( by: chunkBlastSize )
seqKoalaData = Channel
.from(protein)
.splitFasta( by: chunkKoalaSize )
seqIPSData = Channel
.from(protein)
.splitFasta( by: chunkIPSSize )
seqWebData = Channel
.from(protein)
.splitFasta( by: chunkWebSize )
ipscan_properties = file(params.ipscanproperties)
if ( params.debug == "true" || params.debug == true ) {
println("Debugging... only the first $params.debugSize chunks will be processed")
// Diferent parts for different processes.
// TODO: With DSL2 this is far simpler
(seq_file1, seq_file2) = seqData.take(params.debugSize).into(2)
(seq_file_blast) = seqBlastData.take(params.debugSize).into(1)
(seq_file_koala) = seqKoalaData.take(params.debugSize).into(1)
(seq_file_ipscan) = seqIPSData.take(params.debugSize).into(1)
(web_seq_file1, web_seq_file2) = seqWebData.take(params.debugSize).into(2)
testNum = ( params.chunkSize.toInteger() * params.debugSize )
seqTestData = Channel
.from(protein)
.splitFasta(by: testNum)
(seq_test) = seqTestData.take(1).into(1)
} else {
println("Process entire dataset")
(seq_file1, seq_file2) = seqData.into(2)
(seq_file_blast) = seqBlastData.into(1)
(seq_file_koala) = seqKoalaData.into(1)
(seq_file_ipscan) = seqIPSData.into(1)
(web_seq_file1, web_seq_file2) = seqWebData.into(2)
seqTestData = Channel
.from(protein)
// Anything for keeping. This is only kept for coherence
(seq_test) = seqTestData.into(1)
}
if ( params.oboFile == "" || params.oboFile == null ) {
oboFile = downloadURL( "http://www.geneontology.org/ontology/gene_ontology.obo", "gene_ontology.obo" )
} else {
oboFile = params.oboFile
}
// Preprocessing GFF File
if ( gffavail ) {
if ( gffclean ) {
process cleanGFF {
publishDir params.resultPath, mode: 'copy'
label 'gffcheck'
input:
file config_file
output:
file "annot.gff" into gff_file
file "annot.gff.clean.txt" into gff_file_log
"""
# get annot file
export escaped=\$(echo '$baseDir')
export basedirvar=\$(echo '\\\$\\{baseDir\\}')
export basedirvar=\$(echo '\\\$\\{baseDir\\}')
input_file=`perl -lae 'if (\$_=~/gffFile\\s*\\=\\s*[\\x27|\\"](\\S+)[\\x27|\\"]/) { \$base = \$1; \$base=~s/\$ENV{'basedirvar'}/\$ENV{'escaped'}/g; print \$base }' $config_file`
if [ "\${input_file: -3}" == ".gz" ]; then
gunzip -c \$input_file > input_gff
input_file=input_gff
fi
agat_convert_sp_gxf2gxf.pl --gff \$input_file -o annot.gff > annot.gff.clean.txt
"""
}
} else {
process copyGFF {
label 'gffcheck'
input:
file config_file
output:
file "annot.gff" into gff_file
"""
# get annot file
export escaped=\$(echo '$baseDir')
export basedirvar=\$(echo '\\\$\\{baseDir\\}')
input_file=`perl -lae 'if (\$_=~/gffFile\\s*\\=\\s*[\\x27|\\"](\\S+)[\\x27|\\"]/) { \$base = \$1; \$base=~s/\$ENV{'basedirvar'}/\$ENV{'escaped'}/g; print \$base }' $config_file`
if [ "\${input_file: -3}" == ".gz" ]; then
gunzip -c \$input_file > input_gff
input_file=input_gff
fi
cp \$input_file annot.gff
"""
}
}
if ( gffstats ) {
process statsGFF {
publishDir params.resultPath, mode: 'copy'
label 'gffcheck'
input:
file gff_file
output:
file "*.txt" into gff_stats
"""
# Generate Stats
agat_sp_statistics.pl --gff $gff_file > ${gff_file}.stats.txt
"""
}
}
} else {
// Dummy empty GFF
process dummyGFF {
label 'gffcheck'
input:
file config_file
output:
file "annot.gff" into gff_file
"""
# empty annot file
touch annot.gff
"""
}
}
// Blast like processes
// TODO: To change for different aligners
diamond = false
if( params.diamond == "true" || params.diamond == true ) {
diamond = true
}
// BlastAnnotMode
blastAnnotMode = "common"
if( params.blastAnnotMode != "" && params.blastAnnotMode != null ) {
blastAnnotMode = params.blastAnnotMode
}
if ( params.blastFile == "" || params.blastFile == null ){
// program-specific parameters
db_name = file(blastDbPath).name
db_path = file(blastDbPath).parent
// Handling Database formatting
formatdbDetect = "false"
if ( diamond ) {
formatDbFileName = blastDbPath + ".dmnd"
formatDbFile = file(formatDbFileName)
if ( formatDbFile.exists() && formatDbFile.size() > 0 ) {
formatdbDetect = "true"
}
if ( formatdbDetect == "false" ) {
process diamondFormat{
label 'diamond'
output:
file "${db_name}_formatdb.dmnd" into formatdb
"""
diamond makedb --in ${db_path}/${db_name} --db "${db_name}_formatdb"
"""
}
} else {
formatdb = blastDbPath
}
} else {
formatDbDir = file( db_path )
filter = ~/${db_name}.*.phr/
def fcount = 0
formatDbDir.eachFileMatch( filter ) { it ->
fcount = fcount + 1
}
if ( fcount > 0 ) {
formatdbDetect = "true"
}
println( formatdbDetect )
if ( formatdbDetect == "false" ) {
// println( "TUR" )
process blastFormat{
label 'blast'
output:
file "${db_name}.p*" into formatdb
"""
makeblastdb -dbtype prot -in ${db_path}/${db_name} -parse_seqids -out ${db_name}
"""
}
} else {
formatdb = blastDbPath
}
}
if ( diamond == true ) {
process diamond{
label 'diamond'
input:
file seq from seq_file_blast
file formatdb_file from formatdb
output:
file "blastXml${seq}" into (blastXmlResults1, blastXmlResults2, blastXmlResults3)
script:
if ( formatdbDetect == "false" ) {
command = "diamond blastp --db ${formatdb_file} --query $seq --outfmt 5 --threads ${task.cpus} --evalue ${params.evalue} --out blastXml${seq}"
} else {
command = "diamond blastp --db ${db_path}/${db_name} --query $seq --outfmt 5 --threads ${task.cpus} --evalue ${params.evalue} --out blastXml${seq}"
}
command
}
} else {
process blast{
label 'blast'
// publishDir "results", mode: 'copy'
input:
file seq from seq_file_blast
file formatdb_file from formatdb
output:
file "blastXml${seq}" into (blastXmlResults1, blastXmlResults2, blastXmlResults3)
script:
if ( formatdbDetect == "false" ) {
command = "blastp -db ${formatdb_file} -query $seq -num_threads ${task.cpus} -evalue ${params.evalue} -out blastXml${seq} -outfmt 5"
} else {
command = "blastp -db ${db_path}/${db_name} -query $seq -num_threads ${task.cpus} -evalue ${params.evalue} -out blastXml${seq} -outfmt 5"
}
command
}
}
} else {
blastInput=file(params.blastFile)
process convertBlast {
// publishDir "results", mode: 'copy'
input:
file blastFile from blastInput
output:
file("*.xml") into (blastXmlResults1, blastXmlResults2, blastXmlResults3)
"""
hugeBlast2XML.pl -blast $blastFile -n 1000 -out blast.res
"""
}
}
if ( kolist != "" || kolist != null ){
process kofamscan{
label 'kofamscan'
input:
file seq from seq_file_koala
output:
file "koala_${seq}" into koalaResults
"""
exec_annotation --cpu ${task.cpus} -p ${koprofiles} -k ${kolist} -o koala_${seq} $seq
"""
}
process kofam_parse {
input:
file "koala_*" from koalaResults.collect()
output:
file allKoala into ( koala_parsed, koala_parsed2 )
"""
mkdir -p output
processHmmscan2TSV.pl "koala_*" output
cat output/koala_* > allKoala
"""
}
// Replacing keggfile
keggfile = koala_parsed
} else {
if (params.keggFile == "" || params.keggFile == null ) {
println "Please run KEGG KO group annotation on the web server http://www.genome.jp/tools/kaas/"
exit 1
} else {
keggfile = file(params.keggFile)
}
}
// GO retrieval from BLAST results
if (params.gogourl != "") {
process blast_annotator {
label 'blastannotator'
maxForks 3
input:
file blastXml from blastXmlResults2.flatMap()
output:
file "blastAnnot" into ( blast_annotator_results1, blast_annotator_results2 )
"""
blast-annotator.pl -in $blastXml -out blastAnnot --hits $params.gogohits --url $params.gogourl -t $blastAnnotMode -q --format blastxml
"""
}
}
process blastDef {
// publishDir "results", mode: 'copy'
tag "${blastXml}"
input:
file blastXml from blastXmlResults3.flatMap()
output:
file "blastDef_${blastXml}.txt" into blastDef_results
"""
definitionFromBlast.pl -in $blastXml -out blastDef_${blastXml}.txt -format xml -q
"""
}
process ipscn {
label 'ipscan'
if ( workflow.containerEngine == "singularity" ) {
containerOptions "--bind ${ipscandata}:/usr/local/interproscan/data"
} else {
containerOptions "--volume ${ipscandata}:/usr/local/interproscan/data"
}
input:
file seq from seq_file_ipscan
file ("interproscan.properties") from file( ipscan_properties )
output:
file("out_interpro_${seq}") into (ipscn_result1, ipscn_result2)
"""
sed 's/*//g' $seq > tmp4ipscn
interproscan.sh -i tmp4ipscn --goterms --iprlookup --pathways -o out_interpro_${seq} -f TSV -T ${params.ipscantmp}
"""
}
process 'cdSearchHit' {
label 'cdSearch'
if ( ! skip_cdSearch ) {
maxForks 1
}
input:
file seq from web_seq_file1
output:
file("out_hit_${seq}") into ( cdSearch_hit_result1, cdSearch_hit_result2 )
script:
if ( skip_cdSearch ) {
// Dummy content
command = "touch out_hit_${seq}"
} else {
command = "submitCDsearch.pl -o out_hit_${seq} -in $seq"
}
command
}
process 'cdSearchFeat' {
label 'cdSearch'
if ( ! skip_cdSearch ) {
maxForks 1
}
input:
file seq from web_seq_file2
output:
file("out_feat_${seq}") into ( cdSearch_feat_result1, cdSearch_feat_result2 )
script:
if ( skip_cdSearch ) {
// Dummy content
command = "touch out_feat_${seq}"
} else {
command = "submitCDsearch.pl -t feats -o out_feat_${seq} -in $seq"
}
command
}
if ( skip_sigtarp ) {
process 'signalP_dummy' {
input:
file seq from seq_file1
output:
file("out_signalp_${seq}") into (signalP_result1, signalP_result2)
"""
touch out_signalp_${seq}
"""
}
process 'targetP_dummy' {
input:
file seq from seq_file2
output:
file("out_targetp_${seq}") into (targetP_result1, targetP_result2)
"""
touch out_targetp_${seq}
"""
}
} else {
process 'signalP' {
label 'sigtarp'
input:
file seq from seq_file1
output:
file("out_signalp_${seq}") into (signalP_result1, signalP_result2)
"""
signalp -fasta $seq -stdout > out_signalp_${seq}
"""
}
process 'targetP' {
label 'sigtarp'
input:
file seq from seq_file2
output:
file("out_targetp_${seq}") into (targetP_result1, targetP_result2)
"""
targetp -fasta $seq -stdout > out_targetp_${seq}
"""
}
}
// Database setup below
process initDB {
input:
file config_file
file gff_file
file seq from seq_test
output:
file 'config' into (config4perl7, config4perl8, config4perl10)
script:
command = "mkdir -p $params.resultPath\n"
command += "sed 's/^\\s*params\\s*{\\s*\$//gi' $config_file | sed 's/^\\s*}\\s*\$//gi' | sed '/^\\s*\$/d' | sed 's/\\s\\=\\s/:/gi' | sed '/^\\s*\\/\\//d' > configt\n"
command += "export escaped=\$(echo '$baseDir')\n"
command += "export basedirvar=\$(echo '\\\$\\{baseDir\\}')\n"
command += "perl -lae '\$_=~s/\$ENV{'basedirvar'}/\$ENV{'escaped'}/g; print;' configt > config\n"
if ( mysql ) {
// Add dbhost to config
command += "echo \"\$(cat config)\n dbhost:${dbhost}\" > configIn ;\n"
command += "fa_main.v1.pl init -conf configIn"
if ( gffavail && gffclean ) {
command += " -gff ${gff_file}"
}
} else {
if (exists) {
log.info "SQLite database ${dbFileName} exists. We proceed anyway..."
}
command += "fa_main.v1.pl init -conf config"
if ( gffavail && gffclean ) {
command += " -gff ${gff_file}"
}
}
if ( params.debug=="TRUE"||params.debug=="true" ) {
// If in debug mode, we restrict de seq entries we process
command += " -fasta ${seq}"
}
if ( params.rmversion=="TRUE"||params.rmversion=="true" ) {
// If remove versioning in protein sequences (for cases like ENSEMBL)
command += " -rmversion"
}
command
}
if ( ! koentries ) {
process 'kegg_download'{
maxForks 1
input:
file keggfile from keggfile
file config from config4perl8
output:
file("down_kegg") into (down_kegg)
script:
command = "download_kegg_KAAS.pl -input $keggfile -conf $config > done 2>err"
command
}
} else {
process 'kegg_download_dummy' {
input:
file keggfile from keggfile
file config from config4perl8
output:
file("down_kegg") into (down_kegg)
script:
command = "touch down_kegg"
command
}
}
// Data upload process
process 'data_upload' {
maxForks 1
label 'upload'
input:
file "def*" from blastDef_results.collect()
file "out_signalp*" from signalP_result1.collect()
file "out_targetp*" from targetP_result1.collect()