Workflow: Trim Galore RNA-Seq pipeline single-read strand specific
Note: should be updated The original [BioWardrobe's](https://biowardrobe.com) [PubMed ID:26248465](https://www.ncbi.nlm.nih.gov/pubmed/26248465) **RNA-Seq** basic analysis for a **single-end** experiment. A corresponded input [FASTQ](http://maq.sourceforge.net/fastq.shtml) file has to be provided. Current workflow should be used only with the single-end RNA-Seq data. It performs the following steps: 1. Trim adapters from input FASTQ file 2. Use STAR to align reads from input FASTQ file according to the predefined reference indices; generate unsorted BAM file and alignment statistics file 3. Use fastx_quality_stats to analyze input FASTQ file and generate quality statistics file 4. Use samtools sort to generate coordinate sorted BAM(+BAI) file pair from the unsorted BAM file obtained on the step 1 (after running STAR) 5. Generate BigWig file on the base of sorted BAM file 6. Map input FASTQ file to predefined rRNA reference indices using Bowtie to define the level of rRNA contamination; export resulted statistics to file 7. Calculate isoform expression level for the sorted BAM file and GTF/TAB annotation file using GEEP reads-counting utility; export results to file
- Selected
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- Default Values
- Nested Workflows
- Tools
- Inputs/Outputs
Inputs
ID | Type | Title | Doc |
---|---|---|---|
threads | Integer (Optional) | Number of threads |
Number of threads for those steps that support multithreading |
fastq_file | File [FASTQ] | FASTQ input file |
Reads data in a FASTQ format |
clip_3p_end | Integer (Optional) | Clip from 3p end |
Number of bases to clip from the 3p end |
clip_5p_end | Integer (Optional) | Clip from 5p end |
Number of bases to clip from the 5p end |
exclude_chr | String (Optional) | Chromosome to be excluded in rpkm calculation |
Chromosome to be excluded in rpkm calculation |
annotation_file | File [GTF] | Annotation file |
GTF or TAB-separated annotation file |
chrom_length_file | File [Textual format] | Chromosome length file |
Chromosome length file |
star_indices_folder | Directory | STAR indices folder |
Path to STAR generated indices |
bowtie_indices_folder | Directory | BowTie Ribosomal Indices |
Path to Bowtie generated indices |
Steps
ID | Runs | Label | Doc |
---|---|---|---|
rename |
../tools/rename.cwl
(CommandLineTool)
|
Tool renames `source_file` to `target_filename`. Input `target_filename` should be set as string. If it's a full path, only basename will be used. If BAI file is present, it will be renamed too |
|
get_stat |
../tools/collect-statistics-rna-seq.cwl
(CommandLineTool)
|
Tool processes and combines log files generated by Trimgalore, Bowtie, Samtools and MACS2. |
|
trim_fastq |
../tools/trimgalore.cwl
(CommandLineTool)
|
Tool runs Trimgalore - the wrapper around Cutadapt and FastQC to consistently apply adapter and quality trimming
to FastQ files. |
|
star_aligner |
../tools/star-alignreads.cwl
(CommandLineTool)
|
Tool runs STAR alignReads. |
|
extract_fastq |
../tools/extract-fastq.cwl
(CommandLineTool)
|
Tool to decompress input FASTQ file Bash script's logic: - disable case sensitive glob check - check if root name of input file already include '.fastq' or '.fq' extension. If yes, set DEFAULT_EXT to \"\" - check file type, decompress if needed - return 1, if file type is not recognized This script also works of input file doesn't have any extension at all |
|
bowtie_aligner |
../tools/bowtie-alignreads.cwl
(CommandLineTool)
|
Tool maps input raw reads files to reference genome using Bowtie. |
|
group_isoforms |
../tools/group-isoforms.cwl
(CommandLineTool)
|
Tool runs get_gene_n_tss.R script to group isoforms by gene and common TSS |
|
rpkm_calculation |
../tools/geep.cwl
(CommandLineTool)
|
geep |
Tool calculates RPKM values grouped by isoforms or genes. |
get_bam_statistics |
../tools/samtools-stats.cwl
(CommandLineTool)
|
Generates statistics for the input BAM file. |
|
fastx_quality_stats |
../tools/fastx-quality-stats.cwl
(CommandLineTool)
|
Tool calculates statistics on the base of FASTQ file quality scores. If `output_filename` is not provided call function `default_output_filename` to return default output file name generated as `input_file` basename + `.fastxstat` extension. |
|
samtools_sort_index |
../tools/samtools-sort-index.cwl
(CommandLineTool)
|
Tool to sort and index input BAM/SAM/CRAM.
If input `trigger` is set to `true` or isn't set at all (`true` is used by default), run `samtools sort` and
`samtools index`, return sorted BAM and BAI/CSI index file.
If input `trigger` is set to `false`, return unchanged `sort_input` (BAM/SAM/CRAM) and index (BAI/CSI, if provided in
`secondaryFiles`) files, previously staged into output directory. |
|
bam_to_bigwig_upstream |
../tools/bam-bedgraph-bigwig.cwl
(Workflow)
|
Workflow converts input BAM file into bigWig and bedGraph files. |
|
bam_to_bigwig_downstream |
../tools/bam-bedgraph-bigwig.cwl
(Workflow)
|
Workflow converts input BAM file into bigWig and bedGraph files. |
Outputs
ID | Type | Label | Doc |
---|---|---|---|
bowtie_log | File [Textual format] | Bowtie alignment log |
Bowtie alignment log file |
rpkm_genes | File [TSV] | RPKM, grouped by gene name |
Calculated rpkm values, grouped by gene name |
bambai_pair | File [BAM] | Coordinate sorted BAM alignment file (+index BAI) |
Coordinate sorted BAM file and BAI index file |
star_sj_log | File (Optional) [Textual format] | STAR sj log |
STAR SJ.out.tab |
trim_report | File | TrimGalore report |
TrimGalore generated log |
get_stat_log | File (Optional) [YAML] | YAML formatted combined log |
YAML formatted combined log |
star_out_log | File (Optional) [Textual format] | STAR log out |
STAR Log.out |
rpkm_isoforms | File [CSV] | RPKM, grouped by isoforms |
Calculated rpkm values, grouped by isoforms |
star_final_log | File [Textual format] | STAR final log |
STAR Log.final.out |
bigwig_upstream | File [bigWig] | BigWig file |
Generated BigWig file |
rpkm_common_tss | File [TSV] | RPKM, grouped by common TSS |
Calculated rpkm values, grouped by common TSS |
star_stdout_log | File (Optional) [Textual format] | STAR stdout log |
STAR Log.std.out |
fastx_statistics | File [Textual format] | FASTQ statistics |
fastx_quality_stats generated FASTQ file quality statistics file |
bigwig_downstream | File [bigWig] | BigWig file |
Generated BigWig file |
get_stat_markdown | File (Optional) [TIDE TXT] | Markdown formatted combined log |
Markdown formatted combined log |
star_progress_log | File (Optional) [Textual format] | STAR progress log |
STAR Log.progress.out |
get_formatted_stats | File (Optional) [Textual format] | Bowtie, STAR and GEEP mapping stats |
Processed and combined Bowtie & STAR aligner and GEEP logs |
bam_statistics_report | File [Textual format] | BAM statistics report |
BAM statistics report (right after alignment and sorting) |
https://w3id.org/cwl/view/git/5e7385b8cfa4ddae822fff37b6bd22eb0370b389/workflows/trim-rnaseq-se-dutp.cwl