Hello again, when I run for one sample as test with treat file I get this error when the pipeline arrive to the alignment step :

running same command in a local environnement (with Bowtie2/2.4.2) works perfectly. I tryied with several docker images of Bowtie2 and always got same error.
Thanks in advance!

hello, when I use CnR-flow for one sample with treat file but no ctrl file. It detected the input,but said Unknown variable 'R2_Files'.
My command is :
nextflow run CnR-flow --mode run
The output is:
"N E X T F L O W ~ version 20.10.0
Launching RenneLab/CnR-flow [golden_euler] - revision: aa7cf33 [master]

Utilizing Run Mode: run

-- Preparing Workflow Environment --

2 Input Files Detected.

-- Executing Workflow --

[- ] process > CnR_S0_A_GetSeqLen -
[- ] process > CnR_S0_B_MergeFastqs -
[- ] process > CnR_S0_C_FastQCPre -
[- ] process > CnR_S1_A_Trim -
[- ] process > CnR_S1_B_Retrim -
[- ] process > CnR_S1_C_FastQCPost -
[- ] process > CnR_S2_A_Aln_Ref -
[- ] process > CnR_S2_B_Modify_Aln -
Error executing process > 'CnR_S0_B_MergeFastqs (Aged_HSC_CTCF_10)'

Caused by:
Unknown variable 'R2_Files' -- Make sure it is not misspelt and defined somewhere in the script before using it

Source block:
run_id = "${task.tag}.${task.process}"
out_log_name = "${run_id}.nf.log.txt"
merge_fastqs_dir = "${params.merge_fastqs_dir}"
R1_files = fastq.findAll {fn ->
"${fn}".contains("R1") || "${fn}".contains("_1.f") || "${fn}".contains("1")
}
R2_files = fastq.findAll {fn ->
"${fn}".contains("R2") || "${fn}".contains("_2.f") || "${fn}".contains("2")
}
R1_out_file = "${params.merge_fastqs_dir}/${name}_R1_001.fastq.gz"
R2_out_file = "${params.merge_fastqs_dir}/${name}_R2_001.fastq.gz"
if( R1_files.size() < 1 || R2_files.size() < 1 ) {
message = "Merge Error:\nR1 Files: ${R1_files}\nR2 Files: ${R2_Files}"
throw new Exception(message)
}
check_command = ""
if( "${R1_files[0]}".endsWith('.gz') ) {
check_command = "gzip -vt ${R1_files.join(' ')} ${R2_files.join(' ')}"
}
if( R1_files.size() == 1 && R2_files.size() == 1 ) {
command = '''
echo "No Merge Necessary. Renaming Files..."
mkdir !{merge_fastqs_dir}

              # Check File Integrity if gzipped
              !{check_command}

              set -v -H -o history
              mv -v "!{R1_files[0]}" "!{R1_out_file}"
              mv -v "!{R2_files[0]}" "!{R2_out_file}"
              set +v +H +o history
              R1_OUT_LEN=$(zcat -f !{R1_out_file} | wc -l)
              R2_OUT_LEN=$(zcat -f !{R2_out_file} | wc -l)
              echo "R1 Lines: ${R1_OUT_LEN}"
              echo "R2 Lines: ${R2_OUT_LEN}"
              if [ "${R1_OUT_LEN}" == "0" || "${R2_OUT_LEN}" == "0" ]; then
                  echo "Input file of zero length detected."
                  exit 1
              fi
              '''
          } else {
              command = '''
              mkdir !{merge_fastqs_dir}

[- ] process > CnR_S0_A_GetSeqLen [ 0%] 0 of 1
[- ] process > CnR_S0_B_MergeFastqs -
[- ] process > CnR_S0_C_FastQCPre -
[- ] process > CnR_S1_A_Trim -
[- ] process > CnR_S1_B_Retrim -
[- ] process > CnR_S1_C_FastQCPost -
[- ] process > CnR_S2_A_Aln_Ref -
[- ] process > CnR_S2_B_Modify_Aln -
[- ] process > CnR_S2_C_Make_Bdg -
[- ] process > CnR_S3_A_Aln_Spike -
[- ] process > CnR_S3_B_Norm_Bdg -
[- ] process > CnR_S4_A_Make_BigWig -
[- ] process > CnR_S5_A_Peaks_MACS -
[- ] process > CnR_S5_B_Peaks_SEACR -
Error executing process > 'CnR_S0_B_MergeFastqs (Aged_HSC_CTCF_10)'

Caused by:
Unknown variable 'R2_Files' -- Make sure it is not misspelt and defined somewhere in the script before using it

Source block:
run_id = "${task.tag}.${task.process}"
out_log_name = "${run_id}.nf.log.txt"
merge_fastqs_dir = "${params.merge_fastqs_dir}"
R1_files = fastq.findAll {fn ->
"${fn}".contains("R1") || "${fn}".contains("_1.f") || "${fn}".contains("1")
}
R2_files = fastq.findAll {fn ->
"${fn}".contains("R2") || "${fn}".contains("_2.f") || "${fn}".contains("2")
}
R1_out_file = "${params.merge_fastqs_dir}/${name}_R1_001.fastq.gz"
R2_out_file = "${params.merge_fastqs_dir}/${name}_R2_001.fastq.gz"
if( R1_files.size() < 1 || R2_files.size() < 1 ) {
message = "Merge Error:\nR1 Files: ${R1_files}\nR2 Files: ${R2_Files}"
throw new Exception(message)
}
check_command = ""
if( "${R1_files[0]}".endsWith('.gz') ) {
check_command = "gzip -vt ${R1_files.join(' ')} ${R2_files.join(' ')}"
}
if( R1_files.size() == 1 && R2_files.size() == 1 ) {
command = '''
echo "No Merge Necessary. Renaming Files..."
mkdir !{merge_fastqs_dir}

              # Check File Integrity if gzipped
              !{check_command}

              set -v -H -o history
              mv -v "!{R1_files[0]}" "!{R1_out_file}"
              mv -v "!{R2_files[0]}" "!{R2_out_file}"
              set +v +H +o history
              R1_OUT_LEN=$(zcat -f !{R1_out_file} | wc -l)
              R2_OUT_LEN=$(zcat -f !{R2_out_file} | wc -l)
              echo "R1 Lines: ${R1_OUT_LEN}"
              echo "R2 Lines: ${R2_OUT_LEN}"
              if [ "${R1_OUT_LEN}" == "0" || "${R2_OUT_LEN}" == "0" ]; then
                  echo "Input file of zero length detected."
                  exit 1
              fi
              '''
          } else {
              command = '''
              mkdir !{merge_fastqs_dir}
              mkdir !{merge_fastqs_dir}

              # Check File Integrity if gzipped
              !{check_command}

              echo -e "\\nCombining Files: !{R1_files.join(' ')}"
              echo "    Into: !{R1_out_file}"
              set -v -H -o history
              cat '!{R1_files.join("' '")}' > '!{R1_out_file}'
              set +v +H +o history

              echo -e "\\nCombining Files: !{R2_files.join(' ')}"
              echo "    Into: !{R2_out_file}"
              set -v -H -o history
              cat '!{R2_files.join("' '")}' > '!{R2_out_file}'
              set +v +H +o history

              R1_OUT_LEN=$(zcat -f !{R1_out_file} | wc -l)
              R2_OUT_LEN=$(zcat -f !{R2_out_file} | wc -l)
              echo "R1 Lines: ${R1_OUT_LEN}"
              echo "R2 Lines: ${R2_OUT_LEN}"
              if [ "${R1_OUT_LEN}" == "0" || "${R2_OUT_LEN}" == "0" ]; then
                  echo "Input file of zero length detected."
                  exit 1
              fi
              '''
          }

command

Tip: when you have fixed the problem you can continue the execution adding the option -resume to the run command line
"
So what's wrong?
Thanks.

Hello,

Thanks for making this pipeline, I'm excited to give it a try.
After set up the genome fa and fastq file paths, I tried prep_fasta and list_refs , they all worked well, but both dry_run and run seem not to work. Here is it prints:

N E X T F L O W ~ version 20.10.0
Launching RenneLab/CnR-flow [jolly_elion] - revision: fb398b9301 [master]
Utilizing Run Mode: dry_run
-- Preparing Workflow Environment --
74 Input Files Detected.
-- Executing Workflow --

N E X T F L O W ~ version 20.10.0
Launching RenneLab/CnR-flow [special_ride] - revision: fb398b9301 [master]
Utilizing Run Mode: run
-- Preparing Workflow Environment --
74 Input Files Detected.
-- Executing Workflow --

Could you please provide some guidance on where to look for the problem? Thank you!

Best,
Ellie

Hi,

Many thanks for creating such a wonderful workflow for cut and run analysis. I am excited to try this workflow, this is my first attempt of using NextFlow, so perhaps I am missing some steps that are quite obvious. Any pointers would be most appreicated

I am encountering an error while running nextflow run CnR-flow --mode validate_all

My nextflow.config file (.nextflow/assets/RenneLab/CnR-flow/nextflow.config) looks like below.

params {
    // ------- Dependency Configuration --------
    // Configuration using conda is recommended for most systems.
    // Each dependency can only have one type of resource configured:
    // (Ex: bowtie2_module OR bowtie2_conda)

    // Dependency Configuration Using Anaconda
    // Miniconda Install Instructions:
    //     https://docs.conda.io/en/latest/miniconda.html
    //
    // -- External Conda Environments:
    facount_conda          = 'bioconda::ucsc-facount=366'
    bowtie2_conda          = 'bioconda::bowtie2=2.4.1'
    fastqc_conda           = 'bioconda::fastqc=0.11.9'
    trimmomatic_conda      = 'bioconda::trimmomatic=0.39'
    kseqtest_conda         = 'RenneLab::cutruntools-exec'
    bedtools_conda         = 'bioconda::bedtools=2.29.2'
    macs2_conda            = 'bioconda::macs2=2.2.6'
    R_conda                = 'r=3.6.0'
    seacr_conda            = "${params.R_conda} ${params.bedtools_conda}"
    samtools_conda         = 'bioconda::samtools=1.9'
    bedgraphtobigwig_conda = 'conda-forge::libpng conda-forge::libuuid conda-forge::mysql-connector-c conda-forge::openssl conda-forge::zlib bioconda::ucsc-bedgraphtobigwig=377'

My guess is I did not have bowtie2 installed in the conda? So I tried to install it with conda install -c bioconda bowtie2=2.4.1. Checking if the bowtie2 package is available with conda list bowtie2 gave me the result ;

# packages in environment at /home/zfadlullah/anaconda3:
#
# Name                    Version                   Build  Channel
bowtie2                   2.4.1            py38he513fc3_0    bioconda

I think bowtie2 installed, so I am not sure why I am getting the error.

Best,
Zaki

Firstly, I’d like to thank you for this amazing work. I’m trying to apply the pipeline to analyze ChipSeq data. When attempting to run the pipeline in “validate mode” then calling the downloaded images of the system (docker images predefined in “nextflow.config”) with singularity containers, the 'bowtie2' image gives a container string parsing error with exit status (127). When I replaced the version with another updated one, I got the same error with “Samtool” then with “bedgraphtobigwig” and other docker images.

appreciate your time and effort
Many thanks.

System information:
nextflow version : 22.04.5.5708
HPC
Local and Slurm executor were tested
Singularity: 3.8.7-1.el7

Just like to point out a misplaced comma in example file to use fastq_groups:

Example:
// fastq_groups = [
// 'group_1_name': ['treat': 'relpath/to/treat1*R{1,2}*',
// 'ctrl': 'relpath/to/ctrl1*R{1,2}*',
// ]
// 'group_2_name': ['treat': ['relpath/to/g2_treat1*R{1,2}*'
// '/abs/path/to/g2_treat2*R{1,2}*'
// ],
// 'ctrl': 'relpath/to/g2_ctrl1*R{1,2}*'
// ]
// ]

It runs well for multiple groups after moving the comma to the outside of "]"

Example:
// fastq_groups = [
// 'group_1_name': ['treat': 'relpath/to/treat1*R{1,2}*',
// 'ctrl': 'relpath/to/ctrl1*R{1,2}*'
// ],
// 'group_2_name': ['treat': ['relpath/to/g2_treat1*R{1,2}*'
// '/abs/path/to/g2_treat2*R{1,2}*'
// ],
// 'ctrl': 'relpath/to/g2_ctrl1*R{1,2}*'
// ]
// ]
Thanks!