nf-core/rnaseq v3.6 - Platinum Platypus

[3.6] - 2022-03-04

Enhancements & fixes

• nf-core/tools#1415 - Make --outdir a mandatory parameter
• [#734] - Is a vulnerable picard still used ? log4j vulnerability
• [#744] - Auto-detect and raise error if CSI is required for BAM indexing
• [#750] - Optionally ignore R1 / R2 after UMI extraction process
• [#752] - How to set publishing mode for all processes?
• [#753] - Add warning when user provides --transcript_fasta
• [#754] - DESeq2 QC issue linked to --count_col parameter
• [#755] - Rename RSEM_PREPAREREFERENCE_TRANSCRIPTS process
• [#759] - Empty lines in samplesheet.csv cause a crash
• [#769] - Do not run RSeQC tin.py by default

Parameters

Old parameter	New parameter
	--publish_dir_mode
	--umi_discard_read

NB: Parameter has been updated if both old and new parameter information is present.

NB: Parameter has been added if just the new parameter information is present.

NB: Parameter has been removed if new parameter information isn't present.

nf-core/rnaseq v3.5 - Copper Chameleon

[3.5] - 2021-12-17

Enhancements & fixes

• Port pipeline to the updated Nextflow DSL2 syntax adopted on nf-core/modules
  • Removed --publish_dir_mode as it is no longer required for the new syntax
• Bump minimum Nextflow version from 21.04.0 -> 21.10.3
• Updated pipeline template to nf-core/tools 2.2
• [#664] - Conflict of library names for technical replicates
• [#720] - KeyError 'gene_id' in salmon_tx2gene.py
• [#724] - Deal with warnings generated when native NF processes are used
• [#725] - Untar needs --no-same-owner on DNAnexus
• [#727] - Fix transcriptome staging issues on DNAnexus for rsem/prepareference
• [#728] - Add RSeQC TIN.py as a quality metric for the pipeline

nf-core/rnaseq v3.4 - Aluminium Aardvark

[3.4] - 2021-10-05

Enhancements & fixes

• Software version(s) will now be reported for every module imported during a given pipeline execution
• Added python3 shebang to appropriate scripts in bin/ directory
• [#407] - Filter mouse reads from PDX samples
• [#570] - Update SortMeRNA to use SilvaDB 138 (for commercial use)
• [#690] - Error with post-trimmed read 2 sample names from FastQC in MultiQC
• [#693] - Cutadapt version missing from MultiQC report
• [#697] - pipeline_report.{txt,html} missing from pipeline_info directory
• [#705] - Sample sheet error check false positive

Parameters

Old parameter	New parameter
	--bbsplit_fasta_list
	--bbsplit_index
	--save_bbsplit_reads
	--skip_bbsplit

NB: Parameter has been updated if both old and new parameter information is present.
NB: Parameter has been added if just the new parameter information is present.
NB: Parameter has been removed if parameter information isn't present.

Software dependencies

Note, since the pipeline is now using Nextflow DSL2, each process will be run with its own Biocontainer. This means that on occasion it is entirely possible for the pipeline to be using different versions of the same tool. However, the overall software dependency changes compared to the last release have been listed below for reference.

Dependency	Old version	New version
bbmap		38.93
hisat2	2.2.0	2.2.1
picard	2.23.9	2.25.7
salmon	1.4.0	1.5.2
samtools	1.12	1.13
sortmerna	4.2.0	4.3.4
trim-galore	0.6.6	0.6.7

NB: Dependency has been updated if both old and new version information is present.
NB: Dependency has been added if just the new version information is present.
NB: Dependency has been removed if version information isn't present.

nf-core/rnaseq v3.3 - Bronze Bear

[3.3] - 2021-07-29

Enhancements & fixes

• Updated pipeline template to nf-core/tools 2.1
• [#556] - Genome index isn't recreated with --additional_fasta unless --star_index false
• [#668] - Salmon quant with UMI-tools does not work
• [#674] - Launch pipeline regex fails

Software dependencies

Note, since the pipeline is now using Nextflow DSL2, each process will be run with its own Biocontainer. This means that on occasion it is entirely possible for the pipeline to be using different versions of the same tool. However, the overall software dependency changes compared to the last release have been listed below for reference.

Dependency	Old version	New version
samtools	1.10	1.12
stringtie	2.1.4	2.1.7
umi_tools	1.1.1	1.1.2

NB: Dependency has been updated if both old and new version information is present.
NB: Dependency has been added if just the new version information is present.
NB: Dependency has been removed if version information isn't present.

nf-core/rnaseq v3.2 - Copper Flamingo

[3.2] - 2021-06-18

Enhancements & fixes

• Removed workflow to download data from public databases in favour of using nf-core/fetchngs
• Added a stand-alone Python script bin/fastq_dir_to_samplesheet.py to auto-create samplesheet from a directory of FastQ files
• Added docs about overwriting default container definitions to use latest versions e.g. Pangolin
• [#637] - Add --salmon_quant_libtype parameter to provide the --libType option to salmon quantification
• [#645] - Remove trailing slash from params.igenomes_base
• [#649] - DESeq2 fails with only one sample
• [#652] - Results files have incorrect file names
• [nf-core/viralrecon#201] - Conditional include are not expected to work

Parameters

Old parameter	New parameter
--public_data_ids
--skip_sra_fastq_download
	--salmon_quant_libtype

NB: Parameter has been updated if both old and new parameter information is present.
NB: Parameter has been added if just the new parameter information is present.
NB: Parameter has been removed if parameter information isn't present.

nf-core/rnaseq v3.1 - Lead Alligator

[3.1] - 2021-05-13

⚠️ Major enhancements

• Samplesheet format has changed from group,replicate,fastq_1,fastq_2,strandedness to sample,fastq_1,fastq_2,strandedness
  • This gives users the flexibility to name their samples however they wish (see #550)
  • PCA generated by DESeq2 will now be monochrome and will not be grouped by using the replicate id
• Updated Nextflow version to v21.04.0 (see nextflow#572)
• Restructure pipeline scripts into modules/, subworkflows/ and workflows/ directories

Enhancements & fixes

• Updated pipeline template to nf-core/tools 1.14
• Initial implementation of a standardised samplesheet JSON schema to use with user interfaces and for validation
• Only FastQ files that require to be concatenated will be passed to CAT_FASTQ process
• [#449] - --genomeSAindexNbases will now be auto-calculated before building STAR indices
• [#460] - Auto-detect and bypass featureCounts execution if biotype doesn't exist in GTF
• [#544] - Update test-dataset for pipeline
• [#553] - Make tximport output files using all the samples; identified by @j-andrews7
• [#561] - Add gene symbols to merged output; identified by @grst
• [#563] - samplesheet.csv merge error
• [#567] - Update docs to mention trimgalore core usage nuances
• [#568] - --star_index argument is ignored with --aligner star_rsem option
• [#569] - nextflow edge release documentation for running 3.0
• [#575] - Remove duplicated salmon output files
• [#576] - umi_tools dedup : Run before salmon to dedup counts
• [#582] - Generate a separate bigwig tracks for each strand
• [#583] - Samtools error during run requires use of BAM CSI index
• [#585] - Clarify salmon uncertainty for some transcripts
• [#604] - Additional fasta with GENCODE annotation results in biotype error
• [#610] - save R objects as RDS
• [#619] - implicit declaration of the workflow in main
• [#629] - Add and fix EditorConfig linting in entire pipeline
• [nf-core/modules#423] - Replace publish_by_id module option to publish_by_meta
• [nextflow#2060] - Pipeline execution hang when native task fail to be submitted

Parameters

Old parameter	New parameter
--hisat_build_memory	--hisat2_build_memory
--gtf_count_type	--featurecounts_feature_type
--gtf_group_features_type	--featurecounts_group_type
	--bam_csi_index
	--schema_ignore_params
	--show_hidden_params
	--validate_params
--clusterOptions

NB: Parameter has been updated if both old and new parameter information is present.
NB: Parameter has been added if just the new parameter information is present.
NB: Parameter has been removed if parameter information isn't present.

Software dependencies

Note, since the pipeline is now using Nextflow DSL2, each process will be run with its own Biocontainer. This means that on occasion it is entirely possible for the pipeline to be using different versions of the same tool. However, the overall software dependency changes compared to the last release have been listed below for reference.

Dependency	Old version	New version
bedtools	2.29.2	2.30.0
multiqc	1.9	1.10.1
preseq	2.0.3	3.1.2

NB: Dependency has been updated if both old and new version information is present.
NB: Dependency has been added if just the new version information is present.
NB: Dependency has been removed if version information isn't present.

nf-core/rnaseq v3.0 - Silver Shark

[3.0] - 2020-12-15

⚠️ Major enhancements

• You will need to install Nextflow >=20.11.0-edge to run the pipeline. If you are using Singularity, then features introduced in that release now enable the pipeline to directly download Singularity images hosted by Biocontainers as opposed to performing a conversion from Docker images (see #496).
• The previous default of aligning BAM files using STAR and quantifying using featureCounts (--aligner star) has been removed. The new default is to align with STAR and quantify using Salmon (--aligner star_salmon).
  • This decision was made primarily because of the limitations of featureCounts to appropriately quantify gene expression data. Please see Zhao et al., 2015 and Soneson et al., 2015).
• For similar reasons, quantification will not be performed if using --aligner hisat2 due to the lack of an appropriate option to calculate accurate expression estimates from HISAT2 derived genomic alignments.
  • This pipeline option is still available for those who have a preference for the alignment, QC and other types of downstream analysis compatible with the output of HISAT2. No gene-level quantification results will be generated.
  • In a future release we hope to add back quantitation for HISAT2 using different tools.

Enhancements & fixes

• Updated pipeline template to nf-core/tools 1.12.1
• Bumped Nextflow version 20.07.1 -> 20.11.0-edge
• Added UCSC bedClip module to restrict bedGraph file coordinates to chromosome boundaries
• Check if Bioconda and conda-forge channels are set-up correctly when running with -profile conda
• Use rsem-prepare-reference and not gffread to create transcriptome fasta file
• [#494] - Issue running rnaseq v2.0 (DSL2) with test profile
• [#496] - Direct download of Singularity images via HTTPS
• [#498] - Significantly different versions of STAR in star_rsem (2.7.6a) and star (2.6.1d)
• [#499] - Use of salmon counts for DESeq2
• [#500, #509] - Error with AWS batch params
• [#511] - rsem/star index fails with large genome
• [#515] - Add decoy-aware indexing for salmon
• [#516] - Unexpected error [InvocationTargetException]
• [#525] - sra_ids_to_runinfo.py UnicodeEncodeError

Parameters

Old parameter	New parameter
--fc_extra_attributes	--gtf_extra_attributes
--fc_group_features	--gtf_group_features
--fc_count_type	--gtf_count_type
--fc_group_features_type	--gtf_group_features_type
	--singularity_pull_docker_container
--skip_featurecounts

NB: Parameter has been updated if both old and new parameter information is present.
NB: Parameter has been added if just the new parameter information is present.
NB: Parameter has been removed if parameter information isn't present.

Software dependencies

Note, since the pipeline is now using Nextflow DSL2, each process will be run with its own Biocontainer. This means that on occasion it is entirely possible for the pipeline to be using different versions of the same tool. However, the overall software dependency changes compared to the last release have been listed below for reference.

Dependency	Old version	New version
bioconductor-summarizedexperiment	1.18.1	1.20.0
bioconductor-tximeta	1.6.3	1.8.0
picard	2.23.8	2.23.9
requests		2.24.0
salmon	1.3.0	1.4.0
ucsc-bedclip		377
umi_tools	1.0.1	1.1.1

NB: Dependency has been updated if both old and new version information is present.
NB: Dependency has been added if just the new version information is present.
NB: Dependency has been removed if version information isn't present.

nf-core/rnaseq v2.0 - Titanium Tiger

[2.0] - 2020-11-12

Major enhancements

• Pipeline has been re-implemented in Nextflow DSL2
• All software containers are now exclusively obtained from Biocontainers
• Added a separate workflow to download FastQ files via SRA, ENA or GEO ids and to auto-create the input samplesheet (ENA FTP; see --public_data_ids parameter)
• Added and refined a Groovy lib/ of functions that include the automatic rendering of parameters defined in the JSON schema for the help and summary log information
• Replace edgeR with DESeq2 for the generation of PCA and heatmaps (also included in the MultiQC report)
• Creation of bigWig coverage files using BEDTools and bedGraphToBigWig
• [#70] - Added new genome mapping and quantification route with RSEM via the --aligner star_rsem parameter
• [#72] - Samples skipped due to low alignment reported in the MultiQC report
• [#73, #435] - UMI barcode support
• [#91] - Ability to concatenate multiple runs of the same samples via the input samplesheet
• [#123] - The primary input for the pipeline has changed from --reads glob to samplesheet --input. See usage docs.
• [#197] - Samples failing strand-specificity checks reported in the MultiQC report
• [#227] - Removal of ribosomal RNA via SortMeRNA
• [#419] - Add --additional_fasta parameter to provide ERCC spike-ins, transgenes such as GFP or CAR-T as additional sequences to align to

Other enhancements & fixes

• Updated pipeline template to nf-core/tools 1.11
• Optimise MultiQC configuration for faster run-time on huge sample numbers
• Add information about SILVA licensing when removing rRNA to usage.md
• Fixed ansi colours for pipeline summary, added summary logs of alignment results
• [#281] - Add nag to cite the pipeline in summary
• [#302] - Fixed MDS plot axis labels
• [#338] - Add option for turning on/off STAR command line option (--sjdbGTFfile)
• [#344] - Added multi-core TrimGalore support
• [#351] - Fixes missing Qualimap parameter -p
• [#353] - Fixes an issue where MultiQC fails to run with --skip_biotype_qc option
• [#357] - Fixes broken links
• [#362] - Fix error with gzipped annotation file
• [#384] - Changed SortMeRNA reference dbs path to use stable URLs (v4.2.0)
• [#396] - Deterministic mapping for STAR aligner
• [#412] - Fix Qualimap not being passed on correct strand-specificity parameter
• [#413] - Fix STAR unmapped reads not output
• [#434] - Fix typo reported for work-dir
• [#437] - FastQC uses correct number of threads now
• [#440] - Fixed issue where featureCounts process fails when setting --fc_count_type to gene
• [#452] - Fix --gff input bug
• [#345] - Fixes label name in FastQC process
• [#391] - Make publishDir mode configurable
• [#431] - Update AWS GitHub actions workflow with organization level secrets
• [#435] - Fix a bug where gzipped references were not extracted when --additional_fasta was not specified
• [#435] - Fix a bug where merging of RSEM output would fail if only one fastq provided as input
• [#435] - Correct RSEM output name (was saving counts but calling them TPMs; now saving both properly labelled)
• [#436] - Fix a bug where the RSEM reference could not be built
• [#458] - Fix TMP_DIR for process MarkDuplicates and Qualimap

Parameters

Updated

Old parameter	New parameter
--reads	--input
--igenomesIgnore	--igenomes_ignore
--removeRiboRNA	--remove_ribo_rna
--rRNA_database_manifest	--ribo_database_manifest
--save_nonrRNA_reads	--save_non_ribo_reads
--saveAlignedIntermediates	--save_align_intermeds
--saveReference	--save_reference
--saveTrimmed	--save_trimmed
--saveUnaligned	--save_unaligned
--skipAlignment	--skip_alignment
--skipBiotypeQC	--skip_biotype_qc
--skipDupRadar	--skip_dupradar
--skipFastQC	--skip_fastqc
--skipMultiQC	--skip_multiqc
--skipPreseq	--skip_preseq
--skipQC	--skip_qc
--skipQualimap	--skip_qualimap
--skipRseQC	--skip_rseqc
--skipTrimming	--skip_trimming
--stringTieIgnoreGTF	--stringtie_ignore_gtf

Added

• --additional_fasta - FASTA file to concatenate to genome FASTA file e.g. containing spike-in sequences
• --deseq2_vst - Use vst transformation instead of rlog with DESeq2
• --enable_conda - Run this workflow with Conda. You can also use '-profile conda' instead of providing this parameter
• --min_mapped_reads - Minimum percentage of uniquely mapped reads below which samples are removed from further processing
• --multiqc_title - MultiQC report title. Printed as page header, used for filename if not otherwise specified
• --public_data_ids - File containing SRA/ENA/GEO identifiers one per line in order to download their associated FastQ files
• --publish_dir_mode - Method used to save pipeline results to output directory
• --rsem_index - Path to directory or tar.gz archive for pre-built RSEM index
• --rseqc_modules - Specify the RSeQC modules to run
• --save_merged_fastq - Save FastQ files after merging re-sequenced libraries in the results directory
• --save_umi_intermeds - If this option is specified, intermediate FastQ and BAM files produced by UMI-tools are also saved in the results directory
• --skip_bigwig - Skip bigWig file creation
• --skip_deseq2_qc - Skip DESeq2 PCA and heatmap plotting
• --skip_featurecounts - Skip featureCounts
• --skip_markduplicates - Skip picard MarkDuplicates step
• --skip_sra_fastq_download - Only download metadata for public data database ids and don't download the FastQ files
• --skip_stringtie - Skip StringTie
• --star_ignore_sjdbgtf - See #338
• --umitools_bc_pattern - The UMI barcode pattern to use e.g. 'NNNNNN' indicates that the first 6 nucleotides of the read are from the UMI
• --umitools_extract_method - UMI pattern to use. Can be either 'string' (default) or 'regex'
• --with_umi - Enable UMI-based read deduplication

Removed

• --awsqueue can now be provided via nf-core/configs if using AWS
• --awsregion can now be provided via nf-core/configs if using AWS
• --compressedReference now auto-detected
• --markdup_java_options in favour of updating centrally on nf-core/modules
• --project parameter from old NGI template
• --readPaths is not required since these are provided from the input samplesheet
• --sampleLevel not required
• --singleEnd is now auto-detected from the input samplesheet
• --skipEdgeR qc not performed by DESeq2 instead
• --star_memory in favour of updating centrally on nf-core/modules if required
• Strandedness is now specified at the sample-level via the input samplesheet
  • --forwardStranded
  • --reverseStranded
  • --unStranded
  • --pico

Software dependencies

Note, since the pipeline is now using Nextflow DSL2, each process will be run with its own Biocontainer. This means that on occasion it is entirely possible for the pipeline to be using different versions of the same tool. However, the overall software dependency changes compared to the last release have been listed ...

nf-core/rnaseq version 1.4.2

Version 1.4.2

• Minor version release for keeping Git History in sync
• No changes with respect to 1.4.1 on pipeline level

nf-core/rnaseq version 1.4.1

Version 1.4.1

Major novel changes include:

• Update igenomes.config with NCBI GRCh38 and most recent UCSC genomes
• Set autoMounts = true by default for singularity profile

Pipeline enhancements & fixes

• Fixed parameter warnings #316 and 318
• Fixed #307 - Confusing Info Printout about GFF and GTF