Workflow for processing amplicon pool sequencing data with reference.
This workflow allows you to reconstruct a sequence from an amplicon pool using a reference sequence. To run this workflow, you need the reads from the pool library you want to analyse in FASTQ format, separated into two files: forward and reverse. You will also need your reference sequence in FASTA format. This workflow creates a consensus sequence and a metadata file containing the length of the consensus sequence, the number of reads mapped to it, and the average, minimum, and maximum coverage depth. You can also retrieve a file containing unmapped reads.
Inputs
| ID | Name | Description | Type |
|---|---|---|---|
| FASTQ - Forward | FASTQ - Forward | Enter the bank containing the forward reads. |
|
| FASTQ - Reverse | FASTQ - Reverse | Enter the bank containing the reverse reads. |
|
| Filter FASTQ - Maximum size | Filter FASTQ - Maximum size | Maximum reads filtering size. Leave 0 if no filtering. |
|
| Filter FASTQ - Minimum size | Filter FASTQ - Minimum size | Minimum reads filtering size. Leave 0 if no filtering. |
|
| Forward primer | Forward primer | Enter your forward primer in capital letters. |
|
| Reference sequence | Reference sequence | Enter your reference sequence. |
|
| Reverse primer | Reverse primer | Enter your reverse primer in capital letters. |
|
Steps
| ID | Name | Description |
|---|---|---|
| 7 | FastQC | toolshed.g2.bx.psu.edu/repos/devteam/fastqc/fastqc/0.74+galaxy1 |
| 8 | FastQC | toolshed.g2.bx.psu.edu/repos/devteam/fastqc/fastqc/0.74+galaxy1 |
| 9 | Pear | toolshed.g2.bx.psu.edu/repos/iuc/pear/iuc_pear/0.9.6.4 |
| 10 | Zip collections | __ZIP_COLLECTION__ |
| 11 | Create text file | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_text_file_with_recurring_lines/9.5+galaxy2 |
| 12 | Create text file | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_text_file_with_recurring_lines/9.5+galaxy2 |
| 13 | Filter FASTQ | toolshed.g2.bx.psu.edu/repos/devteam/fastq_filter/fastq_filter/1.1.5+galaxy2 |
| 14 | Bowtie2 | toolshed.g2.bx.psu.edu/repos/devteam/bowtie2/bowtie2/2.5.4+galaxy0 |
| 15 | Regex Find And Replace | toolshed.g2.bx.psu.edu/repos/galaxyp/regex_find_replace/regex1/1.0.3 |
| 16 | Regex Find And Replace | toolshed.g2.bx.psu.edu/repos/galaxyp/regex_find_replace/regex1/1.0.3 |
| 17 | FastQC | toolshed.g2.bx.psu.edu/repos/devteam/fastqc/fastqc/0.74+galaxy1 |
| 18 | Bowtie2 | toolshed.g2.bx.psu.edu/repos/devteam/bowtie2/bowtie2/2.5.4+galaxy0 |
| 19 | Bowtie2 | toolshed.g2.bx.psu.edu/repos/devteam/bowtie2/bowtie2/2.5.4+galaxy0 |
| 20 | Samtools view | toolshed.g2.bx.psu.edu/repos/iuc/samtools_view/samtools_view/1.21+galaxy0 |
| 21 | JBrowse2 | toolshed.g2.bx.psu.edu/repos/fubar/jbrowse2/jbrowse2/3.6.5+galaxy0 |
| 22 | Samtools fastx | toolshed.g2.bx.psu.edu/repos/iuc/samtools_fastx/samtools_fastx/1.21+galaxy0 |
| 23 | Samtools idxstats | toolshed.g2.bx.psu.edu/repos/devteam/samtools_idxstats/samtools_idxstats/2.0.7 |
| 24 | Samtools depth | toolshed.g2.bx.psu.edu/repos/iuc/samtools_depth/samtools_depth/1.21+galaxy0 |
| 25 | MEGAHIT | toolshed.g2.bx.psu.edu/repos/iuc/megahit/megahit/1.2.9+galaxy2 |
| 26 | Add line to file | toolshed.g2.bx.psu.edu/repos/bgruening/add_line_to_file/add_line_to_file/0.1.0 |
| 27 | Datamash | toolshed.g2.bx.psu.edu/repos/iuc/datamash_ops/datamash_ops/1.9+galaxy0 |
| 28 | Cutadapt | toolshed.g2.bx.psu.edu/repos/lparsons/cutadapt/cutadapt/5.1+galaxy0 |
| 29 | Select first | Show beginning1 |
| 30 | SeqKit statistics | toolshed.g2.bx.psu.edu/repos/iuc/seqkit_stats/seqkit_stats/2.10.1+galaxy0 |
| 31 | Cut | Cut1 |
| 32 | Select last | Show tail1 |
| 33 | Cut | Cut1 |
| 34 | Paste | Paste1 |
| 35 | Paste | Paste1 |
| 36 | table rename column | toolshed.g2.bx.psu.edu/repos/recetox/table_pandas_rename_column/table_pandas_rename_column/2.2.3+galaxy0 |
Outputs
| ID | Name | Description | Type |
|---|---|---|---|
| fastqc_forward | fastqc_forward | n/a |
|
| fastqc_reverse | fastqc_reverse | n/a |
|
| fastqc_paired | fastqc_paired | n/a |
|
| visualize_consensus | visualize_consensus | n/a |
|
| metadata | metadata | n/a |
|
Version History
Version 1 (earliest) Created 15th Dec 2025 at 14:38 by Molène Mahé
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Created: 15th Dec 2025 at 14:38
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