Explore Workflows

Reverse the lines in a document, then sort those lines.

https://github.com/common-workflow-language/cwl-v1.2.git

Path: tests/revsort.cwl

Branch/Commit ID: e62f99dd79d6cb9c157cceb458f74200da84f6e9

A basic analysis workflow for paired-end ChIP-Seq experiments with a spike-in control. These sequencing library prep methods are chromatin mapping technologies compared to the ChIP-Seq methodology. Its primary benefits include 1) length filtering, 2) a higher signal-to-noise ratio, and 3) built-in normalization for between sample comparisons. This workflow utilizes the tool MACS2 which calls enriched regions in the target sequence data by identifying the top regions by area under a poisson distribution (of the alignment pileup). ### __Inputs__ *General Info (required\*):* - Experiment short name/Alias* - a unique name for the sample (e.g. what was used on tubes while processing it) - Cells* - sample cell type or organism name - Conditions* - experimental condition name - Catalog # - catalog number for cells from vender/supplier - Primary [genome index](https://scidap.com/tutorials/basic/genome-indices) for peak calling* - preprocessed genome index of sample organism for primary alignment and peak calling - Secondary [genome index](https://scidap.com/tutorials/basic/genome-indices) for spike-in normalization* - preprocessed genome index of spike-in organism for secondary alignment (of unaligned reads from primary alignment) and spike-in normalization, default should be E. coli K-12 - FASTQ file for R1* - read 1 file of a pair-end library - FASTQ file for R2* - read 2 file of a pair-end library *Advanced:* - - Number of bases to clip from the 3p end - used by bowtie aligner to trim <int> bases from 3' (right) end of reads - Number of bases to clip from the 5p end - used by bowtie aligner to trim <int> bases from 5' (left) end of reads - Call samtools rmdup to remove duplicates from sorted BAM file? - toggle on/off to remove duplicate reads from analysis - Fragment Length Filter will retain fragments between set base pair (bp) ranges for peak analysis - drop down menu - `default_below_1000` retains fragments <1000 bp - `histones_130_to_300` retains fragments between 130-300 bp - `TF_below_130` retains fragments <130 bp - Max distance (bp) from gene TSS (in both directions) overlapping which the peak will be assigned to the promoter region - default set to `1000` - Max distance (bp) from the promoter (only in upstream directions) overlapping which the peak will be assigned to the upstream region - default set to `20000` - Number of threads for steps that support multithreading - default set to `2` ### __Outputs__ Intermediate and final downloadable outputs include: - IGV with gene, BigWig (raw and normalized), and stringent peak tracks - quality statistics and visualizations for both R1/R2 input FASTQ files - coordinate sorted BAM file with associated BAI file for primary alignment - read pileup/coverage in BigWig format (raw and normalized) - cleaned bed files (containing fragment coordinates), and spike-in normalized peak-called BED files (also includes \"narrow\" and \"broad\" peaks). - stringent peak call bed file with nearest gene annotations per peak ### __Data Analysis Steps__ 1. Trimming the adapters with TrimGalore. - This step is particularly important when the reads are long and the fragments are short - resulting in sequencing adapters at the ends of reads. If adapter is not removed the read will not map. TrimGalore can recognize standard adapters, such as Illumina or Nextera/Tn5 adapters. 2. Generate quality control statistics of trimmed, unmapped sequence data 3. (Optional) Clipping of 5' and/or 3' end by the specified number of bases. 4. Mapping reads to primary genome index with Bowtie. - Only uniquely mapped reads with less than 3 mismatches are used in the downstream analysis. Results are then sorted and indexed. Final outputs are in bam/bai format, which are also used to extrapolate effects of additional sequencing based on library complexity. 5. (Optional) Removal of duplicates (reads/pairs of reads mapping to exactly the same location). - This step is used to remove reads overamplified during amplification of the library. Unfortunately, it may also remove \"good\" reads. We usually do not remove duplicates unless the library is heavily duplicated. 6. Mapping unaligned reads from primary alignment to secondary genome index with Bowtie. - This step is used to obtain the number of reads for normalization, used to scale the read count pileups from the primary alignment used for peak calling. After normalization, sample pileups/peak may then be appropriately compared to one another assuming an equal use of spike-in material during library preparation. 7. Formatting alignment file to account for fragments based on paired-end BAM. - Generates a filtered and normalized bed file to be used as input for peak calling. 8. Call enriched regions using MACS2. - This step called peaks (broad and narrow) using the MACS2 tool with default parameters and no normalization to a control sample. 9. Generation and formatting of output files. - This step collects read, alignment, and peak statistics, as well asgenerates BigWig coverage/pileup files for display on the browser using IGV. The coverage shows the number of fragments that cover each base in the genome both normalized and unnormalized to the calculated spike-in scaling factor.

https://github.com/datirium/workflows.git

Path: workflows/trim-chipseq-pe-spike.cwl

Branch/Commit ID: 46d3d403ddb240d5a8f4f31ab992b6d6a2686745

https://github.com/common-workflow-language/common-workflow-language.git

Path: v1.0/v1.0/step-valuefrom3-wf.cwl

Branch/Commit ID: 17695244222b0301b37cb749fe4a8d89622cd1ad

https://github.com/common-workflow-language/cwl-v1.2.git

Path: tests/env-wf3.cwl

Branch/Commit ID: ea9f8634e41824ac3f81c3dde698d5f0eef54f1b

https://github.com/common-workflow-language/cwltool.git

Path: cwltool/schemas/v1.0/v1.0/count-lines3-wf.cwl

Branch/Commit ID: e9c83739a93fa0b18f8dea2f98b632a9e32725c9

https://github.com/common-workflow-language/cwl-v1.2.git

Path: tests/fail-unconnected.cwl

Branch/Commit ID: 1f3ef888d9ef2306c828065c460c1800604f0de4

# ChIP-Seq basic analysis workflow for single-read data Reads are aligned to the reference genome with [Bowtie](http://bowtie-bio.sourceforge.net/index.shtml). Results are saved as coordinate sorted [BAM](http://samtools.github.io/hts-specs/SAMv1.pdf) alignment and index BAI files. Optionally, PCR duplicates can be removed. To obtain coverage in [bigWig](https://genome.ucsc.edu/goldenpath/help/bigWig.html) format, average fragment length is calculated by [MACS2](https://github.com/taoliu/MACS), and individual reads are extended to this length in the 3’ direction. Areas of enrichment identified by MACS2 are saved in ENCODE [narrow peak](http://genome.ucsc.edu/FAQ/FAQformat.html#format12) or [broad peak](https://genome.ucsc.edu/FAQ/FAQformat.html#format13) formats. Called peaks together with the nearest genes are saved in TSV format. In addition to basic statistics (number of total/mapped/multi-mapped/unmapped/duplicate reads), pipeline generates several quality control measures. Base frequency plots are used to estimate adapter contamination, a frequent occurrence in low-input ChIP-Seq experiments. Expected distinct reads count from [Preseq](http://smithlabresearch.org/software/preseq/) can be used to estimate read redundancy for a given sequencing depth. Average tag density profiles can be used to estimate ChIP enrichment for promoter proximal histone modifications. Use of different parameters for different antibodies (calling broad or narrow peaks) is possible. Additionally, users can elect to use BAM file from another experiment as control for MACS2 peak calling. ## Cite as *Kartashov AV, Barski A. BioWardrobe: an integrated platform for analysis of epigenomics and transcriptomics data. Genome Biol. 2015;16(1):158. Published 2015 Aug 7. [doi:10.1186/s13059-015-0720-3](https://www.ncbi.nlm.nih.gov/pubmed/26248465)* ## Software versions - Bowtie 1.2.0 - Samtools 1.4 - Preseq 2.0 - MACS2 2.1.1.20160309 - Bedtools 2.26.0 - UCSC userApps v358 ## Inputs | ID | Label | Description | Required | Default | Upstream analyses | | ------------------------- | ---------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------------------------- | :------: | ------- | ------------------------------- | | **fastq\_file** | FASTQ file | Single-read sequencing data in FASTQ format (fastq, fq, bzip2, gzip, zip) | + | | | | **indices\_folder** | Genome indices | Directory with the genome indices generated by Bowtie | + | | genome\_indices/bowtie\_indices | | **annotation\_file** | Genome annotation file | Genome annotation file in TSV format | + | | genome\_indices/annotation | | **genome\_size** | Effective genome size | The length of the mappable genome (hs, mm, ce, dm or number, for example 2.7e9) | + | | genome\_indices/genome\_size | | **chrom\_length** | Chromosome lengths file | Chromosome lengths file in TSV format | + | | genome\_indices/chrom\_length | | **broad\_peak** | Call broad peaks | Make MACS2 call broad peaks by linking nearby highly enriched regions | + | | | | **control\_file** | Control ChIP-Seq single-read experiment | Indexed BAM file from the ChIP-Seq single-read experiment to be used as a control for MACS2 peak calling | | Null | control\_file/bambai\_pair | | **exp\_fragment\_size** | Expected fragment size | Expected fragment size for read extenstion towards 3' end if *force\_fragment\_size* was set to True or if calculated by MACS2 fragment size was less that 80 bp | | 150 | | | **force\_fragment\_size** | Force peak calling with expected fragment size | Make MACS2 don't build the shifting model and use expected fragment size for read extenstion towards 3' end | | False | | | **clip\_3p\_end** | Clip from 3' end | Number of base pairs to clip from 3' end | | 0 | | | **clip\_5p\_end** | Clip from 5' end | Number of base pairs to clip from 5' end | | 0 | | | **remove\_duplicates** | Remove PCR duplicates | Remove PCR duplicates from sorted BAM file | | False | | | **threads** | Number of threads | Number of threads for those steps that support multithreading | | 2 | | ## Outputs | ID | Label | Description | Required | Visualization | | ------------------------ | ---------------------------------- | ------------------------------------------------------------------------------------ | :------: | ------------------------------------------------------------------ | | **fastx\_statistics** | FASTQ quality statistics | FASTQ quality statistics in TSV format | + | *Base Frequency* and *Quality Control* plots in *QC Plots* tab | | **bambai\_pair** | Aligned reads | Coordinate sorted BAM alignment and index BAI files | + | *Nucleotide Sequence Alignments* track in *IGV Genome Browser* tab | | **bigwig** | Genome coverage | Genome coverage in bigWig format | + | *Genome Coverage* track in *IGV Genome Browser* tab | | **iaintersect\_result** | Gene annotated peaks | MACS2 peak file annotated with nearby genes | + | *Peak Coordinates* table in *Peak Calling* tab | | **atdp\_result** | Average Tag Density Plot | Average Tag Density Plot file in TSV format | + | *Average Tag Density Plot* in *QC Plots* tab | | **macs2\_called\_peaks** | Called peaks | Called peaks file with 1-based coordinates in XLS format | + | | | **macs2\_narrow\_peaks** | Narrow peaks | Called peaks file in ENCODE narrow peak format | | *Narrow peaks* track in *IGV Genome Browser* tab | | **macs2\_broad\_peaks** | Broad peaks | Called peaks file in ENCODE broad peak format | | *Broad peaks* track in *IGV Genome Browser* tab | | **preseq\_estimates** | Expected Distinct Reads Count Plot | Expected distinct reads count file from Preseq in TSV format | | *Expected Distinct Reads Count Plot* in *QC Plots* tab | | **workflow\_statistics** | Workflow execution statistics | Overall workflow execution statistics from bowtie\_aligner and samtools\_rmdup steps | + | *Overview* tab and experiment's preview | | **bowtie\_log** | Read alignment log | Read alignment log file from Bowtie | + | |

https://github.com/datirium/workflows.git

Path: workflows/chipseq-se.cwl

Branch/Commit ID: 7ced5a5259dbd8b3fc64456beaeffd44f4a24081

Sequence reads are first cleaned from adapters and transformed into fully bisulfite-converted forward (C->T) and reverse read (G->A conversion of the forward strand) versions, before they are aligned to similarly converted versions of the genome (also C->T and G->A converted). Sequence reads that produce a unique best alignment from the four alignment processes against the bisulfite genomes (which are running in parallel) are then compared to the normal genomic sequence and the methylation state of all cytosine positions in the read is inferred. A read is considered to align uniquely if an alignment has a unique best alignment score (as reported by the AS:i field). If a read produces several alignments with the same number of mismatches or with the same alignment score (AS:i field), a read (or a read-pair) is discarded altogether. On the next step we extract the methylation call for every single C analysed. The position of every single C will be written out to a new output file, depending on its context (CpG, CHG or CHH), whereby methylated Cs will be labelled as forward reads (+), non-methylated Cs as reverse reads (-). The output of the methylation extractor is then transformed into a bedGraph and coverage file. The bedGraph counts output is then used to generate a genome-wide cytosine report which reports the number on every single CpG (optionally every single cytosine) in the genome, irrespective of whether it was covered by any reads or not. As this type of report is informative for cytosines on both strands the output may be fairly large (~46mn CpG positions or >1.2bn total cytosine positions in the human genome).

https://github.com/datirium/workflows.git

Path: workflows/bismark-methylation-se.cwl

Branch/Commit ID: 2caa50434966ebdf4b33e5ca689c2e4df32f9058

https://github.com/common-workflow-language/cwl-utils.git

Path: testdata/record-output-wf_v1_2.cwl

Branch/Commit ID: 124a08ce3389eb49066c34a4163cbbed210a0355

https://github.com/common-workflow-language/cwl-utils.git

Path: testdata/step_valuefrom5_wf_with_id_v1_2.cwl

Branch/Commit ID: e78db9870cb744fe36674f43b3223c688e9989e1