added skip_bowtie parameter to make bowtie alignment optional

conf/modules.config

@@ -696,7 +696,7 @@ if(params.run_crosslinking) {
                 path: { "${params.outdir}/04_crosslinks/icountmini_summaries" },
                 mode: "${params.publish_dir_mode}",
                 saveAs: { filename -> filename.equals('versions.yml') ? null : filename },
-                enabled: false
+                enabled: params.skip_bowtie
             ]
         }
 

nextflow.config

@@ -53,7 +53,7 @@ params {
     skip_fastqc        = false
     skip_umi_extract   = true
     skip_trimming      = false
-    skip_transcriptome = true
+
 
     // Output params
     save_reference        = false
@@ -79,6 +79,8 @@ params {
     extra_trimgalore_args      = "--length 10"
 
     // Alignment
+    skip_transcriptome = true
+    skip_bowtie     = false
     save_unaligned = true // Must always be true for unmapped bt to pass to star
     bowtie_params  = "-v 2 -m 100 --norc --best --strata"
     star_params    = "--outFilterMultimapNmax 1 --outFilterMultimapScoreRange 1 --outSAMattributes All --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterType BySJout --alignIntronMin 20 --alignIntronMax 1000000 --outFilterScoreMin 10 --alignEndsType Extend5pOfRead1 --twopassMode Basic"

subworkflows/local/prepare_genome.nf

@@ -43,6 +43,7 @@ workflow PREPARE_GENOME {
     regions_resolved_gtf           // file: .gtf
     skip_filter_gtf                // value: boolean
     skip_transcriptome             // value: boolean
+    skip_bowtie                    // value: boolean
 
     main:
 
@@ -107,18 +108,22 @@ workflow PREPARE_GENOME {
     //
     // MODULES: Uncompress Bowtie index or generate if required
     //
+
     ch_bt_index = Channel.empty()
-    if (ncrna_genome_index) {
-        if (ncrna_genome_index.toString().endsWith(".tar.gz")) {
-            ch_bt_index = UNTAR_BT ( [ [:], ncrna_genome_index ] ).untar
-            ch_versions  = ch_versions.mix(UNTAR_BT.out.versions)
-        } else {
-            ch_bt_index = Channel.of([ [:] , ncrna_genome_index ])
+
+    if (!skip_bowtie) {
+        if (ncrna_genome_index) {
+            if (ncrna_genome_index.toString().endsWith(".tar.gz")) {
+                ch_bt_index = UNTAR_BT ( [ [:], ncrna_genome_index ] ).untar
+                ch_versions  = ch_versions.mix(UNTAR_BT.out.versions)
+            } else {
+                ch_bt_index = Channel.of([ [:] , ncrna_genome_index ])
+            }
+        }
+        else {
+            ch_bt_index = BOWTIE_BUILD ( ch_ncrna_fasta ).index
+            ch_versions = ch_versions.mix(BOWTIE_BUILD.out.versions)
         }
-    }
-    else {
-        ch_bt_index = BOWTIE_BUILD ( ch_ncrna_fasta ).index
-        ch_versions = ch_versions.mix(BOWTIE_BUILD.out.versions)
     }
 
     //

subworkflows/local/rna_align.nf

@@ -33,52 +33,77 @@ workflow RNA_ALIGN {
     gtf                 // channel: [ val(meta), [ gtf ] ]
     fasta               // channel: [ val(meta), [ fasta/fa ]
     skip_transcriptome  // boolean
+    skip_bowtie         // boolean
 
     main:
     ch_versions = Channel.empty()
     //
     // MODULE: Align reads to ncrna genome
     //
-    BOWTIE_ALIGN (
-        fastq,
-        bt_index,
-        true
-    )
-    ch_versions = ch_versions.mix(BOWTIE_ALIGN.out.versions)
 
-    //
-    // SUBWORKFLOW: Sort, index BAM file 
-    //
-    SAMTOOLS_SORT_NCRNA( BOWTIE_ALIGN.out.bam, fasta )
-    SAMTOOLS_INDEX_NCRNA( SAMTOOLS_SORT_NCRNA.out.bam )
+    unmapped_fastq = fastq
+    premapping_log = Channel.empty()
+    premapped_bam = Channel.empty()
+    premapped_bai = Channel.empty()
+
+    premapped_k1_bam = Channel.empty()
+    premapped_k1_bai = Channel.empty()
+
+    if (!skip_bowtie) {
+        BOWTIE_ALIGN (
+            fastq,
+            bt_index,
+            true
+        )
+        ch_versions = ch_versions.mix(BOWTIE_ALIGN.out.versions)
+
+        //
+        // SUBWORKFLOW: Sort, index BAM file 
+        //
+        SAMTOOLS_SORT_NCRNA( BOWTIE_ALIGN.out.bam )
+        SAMTOOLS_INDEX_NCRNA( SAMTOOLS_SORT_NCRNA.out.bam )
 
-    ch_versions = ch_versions.mix(SAMTOOLS_SORT_NCRNA.out.versions)
-    ch_versions = ch_versions.mix(SAMTOOLS_INDEX_NCRNA.out.versions)
+        ch_versions = ch_versions.mix(SAMTOOLS_SORT_NCRNA.out.versions)
+        ch_versions = ch_versions.mix(SAMTOOLS_INDEX_NCRNA.out.versions)
 
-    /*
-    * MODULE: Align reads to smrna genome, here allowing 100 multimappers but only reporting one alignment per multimapped read
-    * so that we can accurately count it in the crosslink summary later
-    */
+        unmapped_fastq = BOWTIE_ALIGN.out.fastq
+        premapping_log = BOWTIE_ALIGN.out.log
+        premapped_bam = SAMTOOLS_SORT_NCRNA.out.bam
+        premapped_bai = SAMTOOLS_INDEX_NCRNA.out.bai
 
-    BOWTIE_ALIGN_K1 (
-        fastq,
-        bt_index,
-        true
-    )
-    ch_versions = ch_versions.mix(BOWTIE_ALIGN_K1.out.versions)
+        /*
+        * MODULE: Align reads to smrna genome, here allowing 100 multimappers but only reporting one alignment per multimapped read
+        * so that we can accurately count it in the crosslink summary later
+        */
+
+        BOWTIE_ALIGN_K1 (
+            fastq,
+            bt_index,
+            true
+        )
+        ch_versions = ch_versions.mix(BOWTIE_ALIGN_K1.out.versions)
 
-    SAMTOOLS_SORT_NCRNA_K1 ( BOWTIE_ALIGN_K1.out.bam, fasta )
-    ch_versions = ch_versions.mix(SAMTOOLS_SORT_NCRNA_K1.out.versions)
+        SAMTOOLS_SORT_NCRNA_K1 ( BOWTIE_ALIGN_K1.out.bam )
+        ch_versions = ch_versions.mix(SAMTOOLS_SORT_NCRNA_K1.out.versions)
+
+        SAMTOOLS_INDEX_NCRNA_K1 ( SAMTOOLS_SORT_NCRNA_K1.out.bam )
+        ch_versions = ch_versions.mix(SAMTOOLS_INDEX_NCRNA_K1.out.versions)
+
+        premapped_k1_bam = SAMTOOLS_SORT_NCRNA_K1.out.bam
+        premapped_k1_bai = SAMTOOLS_INDEX_NCRNA_K1.out.bai  
+
+    }
+
 
-    SAMTOOLS_INDEX_NCRNA_K1 ( SAMTOOLS_SORT_NCRNA_K1.out.bam )
-    ch_versions = ch_versions.mix(SAMTOOLS_INDEX_NCRNA_K1.out.versions)
 
+
+
     //
     // MODULE: Align reads that did not align to the ncrna genome to the primary genome
     //
     if (skip_transcriptome) {
         STAR_ALIGN_GENOME_ONLY (
-            BOWTIE_ALIGN.out.fastq,
+            unmapped_fastq,
             star_index,
             gtf,
             false,
@@ -129,7 +154,7 @@ workflow RNA_ALIGN {
         ch_transcript_multi_bai   = []
     } else {
         STAR_ALIGN_WITH_TRANSCRIPTOME (
-            BOWTIE_ALIGN.out.fastq,
+            unmapped_fastq,
             star_index,
             gtf,
             false,
@@ -210,11 +235,11 @@ workflow RNA_ALIGN {
 
 
     emit:
-    ncrna_bam        = SAMTOOLS_SORT_NCRNA.out.bam      // channel: [ val(meta), [ bam ] ]
-    ncrna_bai        = SAMTOOLS_INDEX_NCRNA.out.bai     // channel: [ val(meta), [ bai ] ]
-    ncrna_log        = BOWTIE_ALIGN.out.log             // channel: [ val(meta), [ txt ] ]
-    ncrna_k1_bam     = SAMTOOLS_SORT_NCRNA_K1.out.bam                  // channel: [ val(meta), [ bam ] ]
-    ncrna_k1_bai     = SAMTOOLS_INDEX_NCRNA_K1.out.bai                 // channel: [ val(meta), [ bai ] ]
+    ncrna_bam        = premapped_bam      // channel: [ val(meta), [ bam ] ]
+    ncrna_bai        = premapped_bai     // channel: [ val(meta), [ bai ] ]
+    ncrna_log        = premapping_log             // channel: [ val(meta), [ txt ] ]
+    ncrna_k1_bam     = premapped_k1_bam                  // channel: [ val(meta), [ bam ] ]
+    ncrna_k1_bai     = premapped_k1_bai                 // channel: [ val(meta), [ bai ] ]
 
     genome_log             = ch_genome_log                    // channel: [ val(meta), [ txt ] ]
     genome_log_final       = ch_genome_log_final              // channel: [ val(meta), [ txt ] ]

tests/nf_test/test/test_only_alignment.nf.test

@@ -54,4 +54,34 @@ nextflow_pipeline {
             assert !(file("$outputDir/02_alignment/genome/PHO92_A_uniqueMapped.bam.bai").exists())
         }
     }
+
+    test("skip_bowtie_alignment") {
+        tag "skip_bowtie_alignment"
+        tag "test"
+        when {
+            params {
+                outdir = "$outputDir"
+                skip_bowtie = true
+            }
+        }
+
+        then {
+            assert workflow.success
+
+            // NCRNA ALIGNMENT
+            assert !(file("$outputDir/02_alignment/ncrna").exists())
+            assert !(file("$outputDir/02_alignment/ncrna/PHO92_A_ncrna.sorted.bam.bai").exists())
+            assert !(file("$outputDir/02_alignment/ncrna/unmapped/PHO92_A_ncrna.unmapped.fastq.gz").exists())
+
+            // icountmini summaries
+            assert file("$outputDir/04_crosslinks/icountmini_summaries/PHO92.summary_gene.tsv").exists()
+            assert file("$outputDir/04_crosslinks/icountmini_summaries/PHO92.summary_subtype.tsv").exists()
+            assert file("$outputDir/04_crosslinks/icountmini_summaries/PHO92.summary_type.tsv").exists()
+
+            assert !file("$outputDir/04_crosslinks/icountmini_summaries/PHO92.summary_gene_premapadjusted.tsv").exists()
+            assert !file("$outputDir/04_crosslinks/icountmini_summaries/PHO92.summary_subtype_premapadjusted.tsv").exists()
+            assert !file("$outputDir/04_crosslinks/icountmini_summaries/PHO92.summary_type_premapadjusted.tsv").exists()
+
+        }
+    }
 }

workflows/clipseq.nf

@@ -228,6 +228,8 @@ workflow CLIPSEQ {
             ch_regions_gtf,
             ch_regions_filt_gtf,
             ch_regions_resolved_gtf,
+            ch_regions_resolved_gtf_genic,
+            params.skip_bowtie,
             params.skip_filter_gtf,
             params.skip_transcriptome
         )
@@ -295,7 +297,8 @@ workflow CLIPSEQ {
             ch_genome_index,
             ch_gtf,
             ch_fasta,
-            params.skip_transcriptome
+            params.skip_transcriptome,
+            params.skip_bowtie
         )
         ch_versions                = ch_versions.mix(RNA_ALIGN.out.versions)
         ch_ncrna_bam               = RNA_ALIGN.out.ncrna_bam