From 600800673a7ff14f4fd0db5fc60043c315ee7ca9 Mon Sep 17 00:00:00 2001 From: DLBPointon Date: Thu, 8 Aug 2024 15:29:03 +0100 Subject: [PATCH] Updates closes #56 and additions to the output.md file --- docs/images/mqc_fastqc_adapter.png | Bin 23458 -> 0 bytes docs/images/mqc_fastqc_counts.png | Bin 33918 -> 0 bytes docs/images/mqc_fastqc_quality.png | Bin 55769 -> 0 bytes docs/output.md | 364 ++++++++++++++++++++++++++-- modules/local/create_btk_dataset.nf | 2 +- modules/local/filter_fasta.nf | 2 +- modules/local/sanger_tol_btk.nf | 17 +- subworkflows/local/yaml_input.nf | 2 +- workflows/ascc.nf | 2 +- 9 files changed, 359 insertions(+), 30 deletions(-) delete mode 100755 docs/images/mqc_fastqc_adapter.png delete mode 100755 docs/images/mqc_fastqc_counts.png delete mode 100755 docs/images/mqc_fastqc_quality.png diff --git a/docs/images/mqc_fastqc_adapter.png b/docs/images/mqc_fastqc_adapter.png deleted file mode 100755 index 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zXwhtT%ei{n#FzEH|C;yZ>+$!u_x#*+`=L8{b9SH^9&27u3G_Gxqxe`L2UJtdxghk z&-wzDFvLvW{chK5u3{n6GSKKy!P&C6w^IFpbD0bcp^A{{2lcLh_DXj@ybtYvc^;(2 M)78&qol`;+0Fu7JivR!s diff --git a/docs/output.md b/docs/output.md index 5a19f95..f3d8a97 100644 --- a/docs/output.md +++ b/docs/output.md @@ -12,46 +12,374 @@ The directories listed below will be created in the results directory after the The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes data using the following steps: -- [FastQC](#fastqc) - Raw read QC -- [MultiQC](#multiqc) - Aggregate report describing results and QC from the whole pipeline +- [YamlInput](#yamlinput) - +- [Validate TaxID](#validate-taxid) - +- [Filter Fasta](#filter-fasta) - +- [GC Content](#gc-content) - +- [Generate Genome](#generate-genome) - +- [Trailing Ns Check](#trailing-ns-check) - +- [Get KMERS profile](#get-kmers-profile) - +- [Extract Tiara Hits](#extract-tiara-hits) - +- [Mito organellar blast](#mito-organellar-blast) - +- [Plastid organellar blast](#plastid-organellar-blast) - +- [Run FCS Adaptor](#run-fcs-adaptor) - +- [Run FCS-GX](#run-fcs-gx) - +- [Pacbio Barcode Check](#pacbio-barcode-check) - +- [Run Read Coverage](#run-read-coverage) - +- [Run Vecscreen](#run-vecscreen) - +- [Run NT Kraken](#run-nt-kraken) - +- [Nucleotide Diamond Blast](#nucleotide-diamond-blast) - +- [Uniprot Diamond Blast](#uniprot-diamond-blast) - +- [Create BTK dataset](#create-btk-dataset) - +- [Autofilter and check assembly](#autofilter-and-check-assembly) - +- [Generate samplesheet](#generate-samplesheet) - +- [Sanger-TOL BTK](#sanger-tol-btk) - +- [Merge BTK datasets](#merge-btk-datasets) - +- [ASCC Merge Tables](#ascc-merge-tables) - - [Pipeline information](#pipeline-information) - Report metrics generated during the workflow execution -### FastQC +### YamlInput
Output files -- `fastqc/` - - `*_fastqc.html`: FastQC report containing quality metrics. - - `*_fastqc.zip`: Zip archive containing the FastQC report, tab-delimited data file and plot images. +- `NA`
-[FastQC](http://www.bioinformatics.babraham.ac.uk/projects/fastqc/) gives general quality metrics about your sequenced reads. It provides information about the quality score distribution across your reads, per base sequence content (%A/T/G/C), adapter contamination and overrepresented sequences. For further reading and documentation see the [FastQC help pages](http://www.bioinformatics.babraham.ac.uk/projects/fastqc/Help/). +YamlInput parses the input yaml into channels for later use in the pipeline. -![MultiQC - FastQC sequence counts plot](images/mqc_fastqc_counts.png) -![MultiQC - FastQC mean quality scores plot](images/mqc_fastqc_quality.png) +### Validate TaxID -![MultiQC - FastQC adapter content plot](images/mqc_fastqc_adapter.png) +
+Output files + +- `NA` + +
+ +Validate TaxID scans through the taxdump to ensure that the input taxid is present in the nxbi taxdump. + + +### Filter Fasta + +
+Output files + +- `filter/` + `*filtered.fasta` - A fasta file that has been filtered for sequences below a given threshold. + +
+ +By default scaffolds above 1.9Gb are removed from the assembly, as scaffolds of this size are unlikely to truely have contamination. There is also the issue that scaffolds larger than this use a significant amount of resources which hinders production environments. + + +### GC Content + +
+Output files + +- `gc/` + `*-GC_CONTENT.txt` - A text file describing the GC content of the input genome. + +
+ +Calculating the GC content of the input genome. + + +### Generate Genome + +
+Output files + +- `generate/` + `*.genome` - An index-like file describing the input genome. + +
+ +An index-like file containing the scaffold and scaffold length of the input genome. + + +### Trailing Ns Check + +
+Output files + +- `trailingns/` + `*_trim_Ns` - A text file containing a report of the Ns found in the genome. + +
+ +A text file containing a report of the Ns found in the genome. + + +### Get KMERS profile + +
+Output files + +- `get/` + `*_KMER_COUNTS.csv` - A csv file containing kmers and their counts. + +
+ +A csv file containing kmers and their counts. + + +### Extract Tiara Hits + +
+Output files + +- `tiara/` + `*.{txt,txt.gz}` - A text file containing classifications of potential contaminants. + `log_*.{txt,txt.gz}` - A log of the tiara run. + `*.{fasta,fasta.gz}` - An output fasta file. + +
+ +Tiara ... + + +### Mito Organellar Blast + +
+Output files + +- `blast/` + `*.tsv` - A tsv file containing potential contaminants. + +
+ +A BlastN based subworkflow used on the input genome to filter potential contaminants from the genome. + + +### Chloro Organellar Blast + +
+Output files + +- `blast/` + `*.tsv` - A tsv file containing potential contaminants. + +
+ +A BlastN based subworkflow used on the input genome to filter potential contaminants from the genome. + + +### Run FCS Adaptor + +
+Output files + +- `fcs/` + `*.fcs_adaptor_report.txt` - A text file containing potential adaptor sequences and locations. + `*.cleaned_sequences.fa.gz` - Cleaned fasta file. + `*.fcs_adaptor.log` - Log of the fcs run. + `*.pipeline_args.yaml` - Arguments to FCS Adaptor + `*.skipped_trims.jsonl` - Skipped sequences + +
+ +FCS Adaptor Identified potential locations of retained adaptor sequences from the sequencing run. + + +### Run FCS-GX + +
+Output files + +- `fcs/` + `*out/*.fcs_gx_report.txt` - A text file containing potential contaminant locations. + `out/*.taxonomy.rpt` - Taxonomy report of the potential contaminants. + +
+ +FCS-GX Identified potential locations of contaminant sequences. + + +### Pacbio Barcode Check + +
+Output files + +- `filter/` + `*_filtered.txt` - Text file of barcodes found in the genome. + +
+ +Uses BlastN to identify where given barcode sequences may be in the genome. + + +### Run Read Coverage + +
+Output files + +- `samtools/` + `*.bam` - Aligned BAM file. + `*_average_coverage.txt` - Text file containing the coverage information for the genome + +
+ +Mapping the read data to the input genome and calculating the average coverage across it. + + +### Run Vecscreen + +
+Output files + +- `summarise/` + `*.vecscreen_contamination` - A text file containing potential vector contaminant locations. + +
+ +Vecscreen identifies vector contamination in the input sequence. + + +### Run NT Kraken + +
+Output files + +- `kraken2/` + `*.classified{.,_}*'` - Fastq file containing classified sequence. + `*.unclassified{.,_}*'` - Fastq file containing unclassified sequence. + `*classifiedreads.txt` - A text file containing a report on reads which have been classified. + `*report.txt` - Report of Kraken2 run. +- `get/` + `*txt` - Text file containing lineage information of the reported meta genomic data. + +
+ +Kraken assigns taxonomic labels to metagenomic DNA sequences and optionally outputs the fastq of these data. + + +### Nucleotide Diamond Blast + +
+Output files + +- `diamond/` + `*.txt` - A text file containing the genomic locations of hits and scores. +- `reformat/` + `*text` - A Reformated text file continaing the full genomic location of hits and scores. +- `convert/` + `*.hits` - A file containing all hits above the cutoff. + +
+ +Diamond Blast is a sequence aligner for translated and protein sequences, here it is used do identify contamination usin the NCBI db + + +### Uniprot Diamond Blast + +
+Output files + +- `diamond/` + `*.txt` - A text file containing the genomic locations of hits and scores. +- `reformat/` + `*text` - A Reformated text file continaing the full genomic location of hits and scores. +- `convert/` + `*.hits` - A file containing all hits above the cutoff. + +
+ +Diamond Blast is a sequence aligner for translated and protein sequences, here it is used do identify contamination usin the Uniprot db + + +### Create BTK dataset + +
+Output files + +- `create/` + `btk_datasets/` - A btk dataset folder containing data compatible with BTK viewer. + `btk_summary_table_full.tsv` - A TSV file summarising the dataset. + +
+ +Create BTK, creates a BTK_dataset folder compatible with BTK viewer. + + +### Autofilter and check assembly + +
+Output files + +- `autofilter/` + `autofiltered.fasta` - The decontaminated input genome. + `ABNORMAL_CHECK.csv` - Combined FCS and Tiara summary of contamination. + `assembly_filtering_removed_sequences.txt` - Sequences deemed contamination and removed from the above assembly. + `fcs-gx_alarm_indicator_file.txt` - Contains text to control the running of Blobtoolkit. + +
+ +Autofilter and check assembly returns a decontaminated genome file as well as summaries of the contamination found. + + +### Generate samplesheet + +
+Output files + +- `generate/` + `*.csv` - A CSV file containing data locations, for use in Blobtoolkit. + +
+ +This produces a CSV containing information on the read data for use in BlobToolKit. + + +### Sanger-TOL BTK + +
+Output files + +- `sanger/` + `*_btk_out/blobtoolkit/${meta.id}*/` - The BTK dataset folder generated by BTK. + `*_btk_out/blobtoolkit/plots/` - The plots for display in BTK Viewer. + `*_btk_out/blobtoolkit/${meta.id}*/summary.json.gz` - The Summary.json file... + `*_btk_out/busco/*` - The BUSCO results returned by BTK. + `*_btk_out/multiqc/*` - The MultiQC results returned by BTK. + `blobtoolkit_pipeline_info` - The pipeline_info folder. + +
+ +Sanger-Tol/BlobToolKit is a Nextflow re-implementation of the snakemake based BlobToolKit pipeline and produces interactive plots used to identify true contamination and seperate sequence from the main assembly. + + +### Merge BTK datasets + +
+Output files + +- `merge/` + `merged_datasets` - A BTK dataset. + `merged_datasets/btk_busco_summary_table_full.tsv` - A TSV file containing a summary of the btk busco results. + +
+ +This module merged the Create_btk_dataset folder with the Sanger-tol BTK dataset to create one unified dataset for use with btk viewer. -> **NB:** The FastQC plots displayed in the MultiQC report shows _untrimmed_ reads. They may contain adapter sequence and potentially regions with low quality. -### MultiQC +### ASCC Merge Tables
Output files -- `multiqc/` - - `multiqc_report.html`: a standalone HTML file that can be viewed in your web browser. - - `multiqc_data/`: directory containing parsed statistics from the different tools used in the pipeline. - - `multiqc_plots/`: directory containing static images from the report in various formats. +- `ascc/` + `*_contamination_check_merged_table.csv` - .... + `*_contamination_check_merged_table_extended.csv` - .... + `*_phylum_counts_and_coverage.csv` - A CSV report containing information on the hits per phylum and the coverage of the hits..
-[MultiQC](http://multiqc.info) is a visualization tool that generates a single HTML report summarising all samples in your project. Most of the pipeline QC results are visualised in the report and further statistics are available in the report data directory. +Merge Tables merged the summary reports from a number of modules inorder to create a single set of reports. -Results generated by MultiQC collate pipeline QC from supported tools e.g. FastQC. The pipeline has special steps which also allow the software versions to be reported in the MultiQC output for future traceability. For more information about how to use MultiQC reports, see . ### Pipeline information diff --git a/modules/local/create_btk_dataset.nf b/modules/local/create_btk_dataset.nf index a88ac0d..bbad73a 100644 --- a/modules/local/create_btk_dataset.nf +++ b/modules/local/create_btk_dataset.nf @@ -26,7 +26,7 @@ process CREATE_BTK_DATASET { output: tuple val(meta), path("btk_datasets"), emit: btk_datasets tuple val(meta), path("btk_summary_table_full.tsv"), emit: create_summary - path "versions.yml", emit: versions + path "versions.yml", emit: versions when: diff --git a/modules/local/filter_fasta.nf b/modules/local/filter_fasta.nf index 20f4f07..2aec22c 100644 --- a/modules/local/filter_fasta.nf +++ b/modules/local/filter_fasta.nf @@ -49,4 +49,4 @@ process FILTER_FASTA { filter_fasta_by_length: \$(filter_fasta_by_length.py -v) END_VERSIONS """ -} \ No newline at end of file +} diff --git a/modules/local/sanger_tol_btk.nf b/modules/local/sanger_tol_btk.nf index f1dd152..3f012f0 100644 --- a/modules/local/sanger_tol_btk.nf +++ b/modules/local/sanger_tol_btk.nf @@ -17,13 +17,13 @@ process SANGER_TOL_BTK { val gca_accession output: - tuple val(meta), path("${meta.id}_btk_out/blobtoolkit/draft"), emit: dataset - path("${meta.id}_btk_out/blobtoolkit/plots"), emit: plots - path("${meta.id}_btk_out/blobtoolkit/draft/summary.json.gz"), emit: summary_json - path("${meta.id}_btk_out/busco"), emit: busco_data - path("${meta.id}_btk_out/multiqc"), emit: multiqc_report - path("blobtoolkit_pipeline_info"), emit: pipeline_info - path "versions.yml", emit: versions + tuple val(meta), path("${meta.id}_btk_out/blobtoolkit/${meta.id}*"), emit: dataset + path("${meta.id}_btk_out/blobtoolkit/plots"), emit: plots + path("${meta.id}_btk_out/blobtoolkit/${meta.id}*/summary.json.gz"), emit: summary_json + path("${meta.id}_btk_out/busco"), emit: busco_data + path("${meta.id}_btk_out/multiqc"), emit: multiqc_report + path("blobtoolkit_pipeline_info"), emit: pipeline_info + path "versions.yml", emit: versions script: def prefix = task.ext.prefix ?: "${meta.id}" @@ -38,7 +38,8 @@ process SANGER_TOL_BTK { // Seems to be an issue where a nested pipeline can't see the files in the same directory // Running realpath gets around this but the files copied into the folder are - // now just wasted space. + // now just wasted space. Should be fixed with using Mahesh's method of nesting but + // this is proving a bit complicated with BTK // outdir should be an arg diff --git a/subworkflows/local/yaml_input.nf b/subworkflows/local/yaml_input.nf index 6d21ea5..935e503 100644 --- a/subworkflows/local/yaml_input.nf +++ b/subworkflows/local/yaml_input.nf @@ -148,7 +148,7 @@ workflow YAML_INPUT { ncbi_taxonomy_path = group.ncbi_taxonomy_path ncbi_rankedlineage_path = group.ncbi_rankedlineage_path busco_lineages_folder = group.busco_lineages_folder - busco_lineages = group.busco_lineages + busco_lineages = group.busco_lineages fcs_gx_database_path = group.fcs_gx_database_path diamond_uniprot_database_path = group.diamond_uniprot_database_path diamond_nr_database_path = group.diamond_nr_database_path diff --git a/workflows/ascc.nf b/workflows/ascc.nf index 9785dba..48b4dc8 100644 --- a/workflows/ascc.nf +++ b/workflows/ascc.nf @@ -326,7 +326,7 @@ workflow ASCC { } .collect() .set { - ch_barcode + ch_barcode // Not in use } ch_versions = ch_versions.mix(PACBIO_BARCODE_CHECK.out.versions)