How To emit nextflow subworkflow outputs ?!

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1

Hi Folks , I Am trying to write nextflow subworkflow : for :
1- Trimming
2- Aligninig trimmed Reads
And unfortunately I didn’t know how to emit Trimmed reads which derived from sub-workflow n°1 to be used into the next sub-workflow n°2 ( Alignment )
Any Advice !
main.nf
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Trimmming Subworkflow

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image763×428 44 KB

Here my Triming subworkflowoutputs :point_down:

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Alignment Subworkflow

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image875×496 41 KB

ERROR
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2

@mribeirodantas what do suggest please :thinking:

3

You have to call the TRIMMED subworkflow so that there’s an output channel to be consumed by the Alignment process. It’s what the error message is saying. Shouldn’t you have trimmedReads where you typed TRIMMED.out.trimmedR?

4

Yes I already create that channel but . Alignment subworkfkow don’t recognise it
I want to align Trim.outputs ( process under TRIMMD Subworkfkow)

TRIMMED SUBWORKFKOW

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image763×428 44 KB

Alignment subworkfkow

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image875×496 41 KB

5

Pictures aren’t the best way to share code. Please, share code blocks so I can know if these are all in the same file, for example. In the code you showed, I don’t see the subworkflows being called.

6

@mribeirodantas No they are not in the same file, They are organised like that :
├── data
│ ├── SRR519926_1.fastq
│ ├── SRR519926_2.fastq
│ ├── SRR519930_1.fastq
│ └── SRR519930_2.fastq
├── main.nf
├── modules
│ ├── Alignment.nf
│ ├── bwa_index.nf
│ ├── FASTQC.nf
│ └── TRIM.nf
├── Ref_genome
│ └── Ecoli.fa
├── samplemain.csv
└── subworkflows
├── alignmentsub.nf
├── RawQC_sub.nf
└── Trim&QC_Sub.nf
Main.nf :point_down:

#!/usr/bin/env nextflow



// Params 
params.reads = "$projectDir/samplemain.csv"
params.reference = "$projectDir/Ref_genome/*.{fasta,fa}"
params.outdir ="$projectDir/outdir"
params.cpus = 3



//Channels

reads = Channel.fromPath(params.reads, checkIfExists: true)
               .splitCsv(header: true)
               .map { row -> tuple(row.sampleId, file(row.read1), file(row.read2)) } 


Channel.fromPath(params.reference)
       .first()
       .set {ref_file}       

      
// Sub-Workflows 

// Raw Reads QC :: nextflow 

include { RAW_QC } from "./subworkflows/RawQC_sub.nf" 

workflow Raw {
 	RAW_QC (reads) 
 	} 

// Trimming +  QC check 

include { TRIMMED } from "./subworkflows/Trim&QC_Sub.nf"  

workflow Trim { 
	TRIMMED (reads) 
	}          
    
// Mapping to Reference 
 
include { ALIGNMENT } from "./subworkflows/alignmentsub.nf"
 
workflow Align { 
	ALIGNMENT (ref_file,TRIMMED)
	}

Trimming Sub-Workflow

/*
========================================================================================
   Trimming Sub-Workflow
========================================================================================
*/


include {Trimming} from '../modules/TRIM.nf'            // Check raw reads Quality 
include {QC_Trim} from '../modules/TRIM.nf'        // Multiqc for raw reads
include {Multiqc_Trim} from '../modules/TRIM.nf'        // Multiqc for raw reads

reads = Channel.fromPath(params.reads, checkIfExists: true)
               .splitCsv(header: true)
               .map { row -> tuple(row.sampleId, file(row.read1), file(row.read2)) } 

workflow TRIMMED {

    take:
        reads

    main:
       Trimming(reads)    
       QC_Trim(Trimming.out.triMmed)  
       Multiqc_Trim(QC_Trim.out.collect()) 
    
   emit: 
      trimmedR = Trimming.out.triMmed.collect()
      trimmedR.view()
}

Alignment Sub-Workflow

/*
========================================================================================
  Alignment Sub-Workflow
========================================================================================
*/


include {index		} from '../modules/bwa_index.nf'          // Index the reference
include {Alignment	} from '../modules/Alignment.nf'         // Alignment based-reference 
include {TRIMMED	} from '../subworkflows/Trim&QC_Sub.nf' // Trimming subworkflow 
  
  
/* channels */

Channel.fromPath(params.reference)
    .first()
    .set { ref_file }
    


workflow ALIGNMENT { 
    take:
        ref
        trimmedReads
    main :
    index(ref_file)
    Alignment(ref_file,index.out.indexes,TRIMMED.out.trimmedR)

}

ERROR

nextflow run main.nf -entry Align
N E X T F L O W  ~  version 23.04.1
Launching `main.nf` [pedantic_jang] DSL2 - revision: 8bc364eb64
Access to 'TRIMMED.out' is undefined since the workflow 'TRIMMED' has not been invoked before accessing the output attribute

 -- Check script './subworkflows/alignmentsub.nf' at line: 27 or see '.nextflow.log' file for more details
7

trimmedReads isn’t a variable you can access everywhere, you need to use the output of TRIMMED in the input of ALIGNMENT. Here I have combined you examples to demonstrate this. See the anonymous workflow at the bottom:

include {Trimming    } from '../modules/TRIM.nf'            // Check raw reads Quality 
include {QC_Trim     } from '../modules/TRIM.nf'            // Multiqc for raw reads
include {Multiqc_Trim} from '../modules/TRIM.nf'            // Multiqc for raw reads
include {index		 } from '../modules/bwa_index.nf'       // Index the reference
include {Alignment	 } from '../modules/Alignment.nf'       // Alignment based-reference 
include {TRIMMED	 } from '../subworkflows/Trim&QC_Sub.nf' // Trimming subworkflow 
  


workflow TRIMMED {

    take:
        reads

    main:
       Trimming(reads)    
       QC_Trim(Trimming.out.triMmed)  
       Multiqc_Trim(QC_Trim.out.collect()) 
    
   emit: 
      trimmedR = Trimming.out.triMmed.collect()
}

workflow ALIGNMENT { 
    take:
        ref
        trimmedReads
    main:
        index(ref_file)
        Alignment(ref_file,index.out.indexes,TRIMMED.out.trimmedR)

}

workflow {
    reads = Channel.fromPath(params.reads, checkIfExists: true)
               .splitCsv(header: true)
               .map { row -> tuple(row.sampleId, file(row.read1), file(row.read2)) } 

    ref_file = Channel.fromPath(params.reference)
                .first()

    TRIMMED(reads)
    ALIGNMENT(ref_file, TRIMMED.out.trimmedR)
}