mapping/star_align

Description

Align fastq files using STAR.

Type

python_script

Contributors

Run this component

Run the following command to execute this component with Nextflow:

cat > params.yaml <<'EOM'  
output: "$id.$key.output./path/to/foo"  
id: "run"  
publish_dir: "output/"  
EOM

nextflow run https://packages.viash-hub.com/vsh/openpipeline.git \  
  -revision 1.0.4 \  
  -main-script target/nextflow/mapping/star_align/main.nf \  
  -params-file params.yaml

Input/Output

Name	Type & Properties	Description
--input --readFilesIn	file required multiple	The FASTQ files to be analyzed. Corresponds to the --readFilesIn argument in the STAR command.
--reference --genomeDir	file required	Path to the reference built by star_build_reference. Corresponds to the --genomeDir argument in the STAR command.
--output --outFileNamePrefix	file required output	Path to output directory. Corresponds to the --outFileNamePrefix argument in the STAR command.

Name

Type & Properties

--input

--readFilesIn

file

required

multiple

--reference

--genomeDir

file

required

--output

--outFileNamePrefix

file

required

output

Run Parameters

Name	Type & Properties	Description
--runRNGseed	integer	random number generator seed.

Name

Type & Properties

--runRNGseed

integer

Genome Parameters

Name	Type & Properties	Description
--genomeLoad	string	mode of shared memory usage for the genome files. Only used with --runMode alignReads. - LoadAndKeep ... load genome into shared and keep it in memory after run - LoadAndRemove ... load genome into shared but remove it after run - LoadAndExit ... load genome into shared memory and exit, keeping the genome in memory for future runs - Remove ... do not map anything, just remove loaded genome from memory - NoSharedMemory ... do not use shared memory, each job will have its own private copy of the genome
--genomeFastaFiles	file multiple	path(s) to the fasta files with the genome sequences, separated by spaces. These files should be plain text FASTA files, they cannot be zipped. Required for the genome generation (--runMode genomeGenerate). Can also be used in the mapping (--runMode alignReads) to add extra (new) sequences to the genome (e.g. spike-ins).
--genomeFileSizes	integer multiple	genome files exact sizes in bytes. Typically, this should not be defined by the user.
--genomeTransformOutput	string multiple	which output to transform back to original genome - SAM ... SAM/BAM alignments - SJ ... splice junctions (SJ.out.tab) - None ... no transformation of the output
--genomeChrSetMitochondrial	string multiple	names of the mitochondrial chromosomes. Presently only used for STARsolo statistics output/

Name

Type & Properties

--genomeLoad

string

--genomeFastaFiles

file

multiple

--genomeFileSizes

integer

multiple

--genomeTransformOutput

string

multiple

--genomeChrSetMitochondrial

string

multiple

Splice Junctions Database

Name	Type & Properties	Description
--sjdbFileChrStartEnd	string multiple	path to the files with genomic coordinates (chr <tab> start <tab> end <tab> strand) for the splice junction introns. Multiple files can be supplied and will be concatenated.
--sjdbGTFfile	file	path to the GTF file with annotations
--sjdbGTFchrPrefix	string	prefix for chromosome names in a GTF file (e.g. 'chr' for using ENSMEBL annotations with UCSC genomes)
--sjdbGTFfeatureExon	string	feature type in GTF file to be used as exons for building transcripts
--sjdbGTFtagExonParentTranscript	string	GTF attribute name for parent transcript ID (default "transcript_id" works for GTF files)
--sjdbGTFtagExonParentGene	string	GTF attribute name for parent gene ID (default "gene_id" works for GTF files)
--sjdbGTFtagExonParentGeneName	string multiple	GTF attribute name for parent gene name
--sjdbGTFtagExonParentGeneType	string multiple	GTF attribute name for parent gene type
--sjdbOverhang	integer	length of the donor/acceptor sequence on each side of the junctions, ideally = (mate_length - 1)
--sjdbScore	integer	extra alignment score for alignments that cross database junctions
--sjdbInsertSave	string	which files to save when sjdb junctions are inserted on the fly at the mapping step - Basic ... only small junction / transcript files - All ... all files including big Genome, SA and SAindex - this will create a complete genome directory

Name

Type & Properties

--sjdbFileChrStartEnd

string

multiple

--sjdbGTFfile

file

--sjdbGTFchrPrefix

string

--sjdbGTFfeatureExon

string

--sjdbGTFtagExonParentTranscript

string

--sjdbGTFtagExonParentGene

string

--sjdbGTFtagExonParentGeneName

string

multiple

--sjdbGTFtagExonParentGeneType

string

multiple

--sjdbOverhang

integer

--sjdbScore

integer

--sjdbInsertSave

string

Variation parameters

Name	Type & Properties	Description
--varVCFfile	string	path to the VCF file that contains variation data. The 10th column should contain the genotype information, e.g. 0/1

Name

Type & Properties

--varVCFfile

string

Read Parameters

Name	Type & Properties	Description
--readFilesType	string	format of input read files - Fastx ... FASTA or FASTQ - SAM SE ... SAM or BAM single-end reads; for BAM use --readFilesCommand samtools view - SAM PE ... SAM or BAM paired-end reads; for BAM use --readFilesCommand samtools view
--readFilesSAMattrKeep	string multiple	for --readFilesType SAM SE/PE, which SAM tags to keep in the output BAM, e.g.: --readFilesSAMtagsKeep RG PL - All ... keep all tags - None ... do not keep any tags
--readFilesManifest	file	path to the "manifest" file with the names of read files. The manifest file should contain 3 tab-separated columns: paired-end reads: read1_file_name $tab$ read2_file_name $tab$ read_group_line. single-end reads: read1_file_name $tab$ - $tab$ read_group_line. Spaces, but not tabs are allowed in file names. If read_group_line does not start with ID:, it can only contain one ID field, and ID: will be added to it. If read_group_line starts with ID:, it can contain several fields separated by $tab$, and all fields will be be copied verbatim into SAM @RG header line.
--readFilesPrefix	string	prefix for the read files names, i.e. it will be added in front of the strings in --readFilesIn
--readFilesCommand	string multiple	command line to execute for each of the input file. This command should generate FASTA or FASTQ text and send it to stdout For example: zcat - to uncompress .gz files, bzcat - to uncompress .bz2 files, etc.
--readMapNumber	integer	number of reads to map from the beginning of the file -1: map all reads
--readMatesLengthsIn	string	Equal/NotEqual - lengths of names,sequences,qualities for both mates are the same / not the same. NotEqual is safe in all situations.
--readNameSeparator	string multiple	character(s) separating the part of the read names that will be trimmed in output (read name after space is always trimmed)
--readQualityScoreBase	integer	number to be subtracted from the ASCII code to get Phred quality score

Name

Type & Properties

--readFilesType

string

--readFilesSAMattrKeep

string

multiple

--readFilesManifest

file

--readFilesPrefix

string

--readFilesCommand

string

multiple

--readMapNumber

integer

--readMatesLengthsIn

string

--readNameSeparator

string

multiple

--readQualityScoreBase

integer

Read Clipping

Name	Type & Properties	Description
--clipAdapterType	string	adapter clipping type - Hamming ... adapter clipping based on Hamming distance, with the number of mismatches controlled by --clip5pAdapterMMp - CellRanger4 ... 5p and 3p adapter clipping similar to CellRanger4. Utilizes Opal package by Martin Sosic: https://github.com/Martinsos/opal - None ... no adapter clipping, all other clip* parameters are disregarded
--clip3pNbases	integer multiple	number(s) of bases to clip from 3p of each mate. If one value is given, it will be assumed the same for both mates.
--clip3pAdapterSeq	string multiple	adapter sequences to clip from 3p of each mate. If one value is given, it will be assumed the same for both mates. - polyA ... polyA sequence with the length equal to read length
--clip3pAdapterMMp	double multiple	max proportion of mismatches for 3p adapter clipping for each mate. If one value is given, it will be assumed the same for both mates.
--clip3pAfterAdapterNbases	integer multiple	number of bases to clip from 3p of each mate after the adapter clipping. If one value is given, it will be assumed the same for both mates.
--clip5pNbases	integer multiple	number(s) of bases to clip from 5p of each mate. If one value is given, it will be assumed the same for both mates.

Name

Type & Properties

--clipAdapterType

string

--clip3pNbases

integer

multiple

--clip3pAdapterSeq

string

multiple

--clip3pAdapterMMp

double

multiple

--clip3pAfterAdapterNbases

integer

multiple

--clip5pNbases

integer

multiple

Limits

Name	Type & Properties	Description
--limitGenomeGenerateRAM	long	maximum available RAM (bytes) for genome generation
--limitIObufferSize	long multiple	max available buffers size (bytes) for input/output, per thread
--limitOutSAMoneReadBytes	long	max size of the SAM record (bytes) for one read. Recommended value: >(2(LengthMate1+LengthMate2+100)outFilterMultimapNmax
--limitOutSJoneRead	integer	max number of junctions for one read (including all multi-mappers)
--limitOutSJcollapsed	integer	max number of collapsed junctions
--limitBAMsortRAM	long	maximum available RAM (bytes) for sorting BAM. If =0, it will be set to the genome index size. 0 value can only be used with --genomeLoad NoSharedMemory option.
--limitSjdbInsertNsj	integer	maximum number of junctions to be inserted to the genome on the fly at the mapping stage, including those from annotations and those detected in the 1st step of the 2-pass run
--limitNreadsSoft	integer	soft limit on the number of reads

Name

Type & Properties

--limitGenomeGenerateRAM

long

--limitIObufferSize

long

multiple

--limitOutSAMoneReadBytes

long

--limitOutSJoneRead

integer

--limitOutSJcollapsed

integer

--limitBAMsortRAM

long

--limitSjdbInsertNsj

integer

--limitNreadsSoft

integer

Output: general

Name	Type & Properties	Description
--outTmpKeep	string	whether to keep the temporary files after STAR runs is finished - None ... remove all temporary files - All ... keep all files
--outStd	string	which output will be directed to stdout (standard out) - Log ... log messages - SAM ... alignments in SAM format (which normally are output to Aligned.out.sam file), normal standard output will go into Log.std.out - BAM_Unsorted ... alignments in BAM format, unsorted. Requires --outSAMtype BAM Unsorted - BAM_SortedByCoordinate ... alignments in BAM format, sorted by coordinate. Requires --outSAMtype BAM SortedByCoordinate - BAM_Quant ... alignments to transcriptome in BAM format, unsorted. Requires --quantMode TranscriptomeSAM
--outReadsUnmapped	string	output of unmapped and partially mapped (i.e. mapped only one mate of a paired end read) reads in separate file(s). - None ... no output - Fastx ... output in separate fasta/fastq files, Unmapped.out.mate1/2
--outQSconversionAdd	integer	add this number to the quality score (e.g. to convert from Illumina to Sanger, use -31)
--outMultimapperOrder	string	order of multimapping alignments in the output files - Old_2.4 ... quasi-random order used before 2.5.0 - Random ... random order of alignments for each multi-mapper. Read mates (pairs) are always adjacent, all alignment for each read stay together. This option will become default in the future releases.

Name

Type & Properties

--outTmpKeep

string

--outStd

string

--outReadsUnmapped

string

--outQSconversionAdd

integer

--outMultimapperOrder

string

Output: SAM and BAM

Name	Type & Properties	Description
--outSAMtype	string multiple	type of SAM/BAM output 1st word: - BAM ... output BAM without sorting - SAM ... output SAM without sorting - None ... no SAM/BAM output 2nd, 3rd: - Unsorted ... standard unsorted - SortedByCoordinate ... sorted by coordinate. This option will allocate extra memory for sorting which can be specified by --limitBAMsortRAM.
--outSAMmode	string	mode of SAM output - None ... no SAM output - Full ... full SAM output - NoQS ... full SAM but without quality scores
--outSAMstrandField	string	Cufflinks-like strand field flag - None ... not used - intronMotif ... strand derived from the intron motif. This option changes the output alignments: reads with inconsistent and/or non-canonical introns are filtered out.
--outSAMattributes	string multiple	a string of desired SAM attributes, in the order desired for the output SAM. Tags can be listed in any combination/order. *Presets: - None ... no attributes - Standard ... NH HI AS nM - All ... NH HI AS nM NM MD jM jI MC ch Alignment: - NH ... number of loci the reads maps to: =1 for unique mappers, >1 for multimappers. Standard SAM tag. - HI ... multiple alignment index, starts with --outSAMattrIHstart (=1 by default). Standard SAM tag. - AS ... local alignment score, +1/-1 for matches/mismateches, score penalties for indels and gaps. For PE reads, total score for two mates. Stadnard SAM tag. - nM ... number of mismatches. For PE reads, sum over two mates. - NM ... edit distance to the reference (number of mismatched + inserted + deleted bases) for each mate. Standard SAM tag. - MD ... string encoding mismatched and deleted reference bases (see standard SAM specifications). Standard SAM tag. - jM ... intron motifs for all junctions (i.e. N in CIGAR): 0: non-canonical; 1: GT/AG, 2: CT/AC, 3: GC/AG, 4: CT/GC, 5: AT/AC, 6: GT/AT. If splice junctions database is used, and a junction is annotated, 20 is added to its motif value. - jI ... start and end of introns for all junctions (1-based). - XS ... alignment strand according to --outSAMstrandField. - MC ... mate's CIGAR string. Standard SAM tag. - ch ... marks all segment of all chimeric alingments for --chimOutType WithinBAM output. - cN ... number of bases clipped from the read ends: 5' and 3' *Variation: - vA ... variant allele - vG ... genomic coordinate of the variant overlapped by the read. - vW ... 1 - alignment passes WASP filtering; 2,3,4,5,6,7 - alignment does not pass WASP filtering. Requires --waspOutputMode SAMtag. STARsolo: - CR CY UR UY ... sequences and quality scores of cell barcodes and UMIs for the solo demultiplexing. - GX GN ... gene ID and gene name for unique-gene reads. - gx gn ... gene IDs and gene names for unique- and multi-gene reads. - CB UB ... error-corrected cell barcodes and UMIs for solo* demultiplexing. Requires --outSAMtype BAM SortedByCoordinate. - sM ... assessment of CB and UMI. - sS ... sequence of the entire barcode (CB,UMI,adapter). - sQ ... quality of the entire barcode. ***Unsupported/undocumented: - ha ... haplotype (1/2) when mapping to the diploid genome. Requires genome generated with --genomeTransformType Diploid . - rB ... alignment block read/genomic coordinates. - vR ... read coordinate of the variant.
--outSAMattrIHstart	integer	start value for the IH attribute. 0 may be required by some downstream software, such as Cufflinks or StringTie.
--outSAMunmapped	string multiple	output of unmapped reads in the SAM format 1st word: - None ... no output - Within ... output unmapped reads within the main SAM file (i.e. Aligned.out.sam) 2nd word: - KeepPairs ... record unmapped mate for each alignment, and, in case of unsorted output, keep it adjacent to its mapped mate. Only affects multi-mapping reads.
--outSAMorder	string	type of sorting for the SAM output Paired: one mate after the other for all paired alignments PairedKeepInputOrder: one mate after the other for all paired alignments, the order is kept the same as in the input FASTQ files
--outSAMprimaryFlag	string	which alignments are considered primary - all others will be marked with 0x100 bit in the FLAG - OneBestScore ... only one alignment with the best score is primary - AllBestScore ... all alignments with the best score are primary
--outSAMreadID	string	read ID record type - Standard ... first word (until space) from the FASTx read ID line, removing /1,/2 from the end - Number ... read number (index) in the FASTx file
--outSAMmapqUnique	integer	0 to 255: the MAPQ value for unique mappers
--outSAMflagOR	integer	0 to 65535: sam FLAG will be bitwise OR'd with this value, i.e. FLAG=FLAG \| outSAMflagOR. This is applied after all flags have been set by STAR, and after outSAMflagAND. Can be used to set specific bits that are not set otherwise.
--outSAMflagAND	integer	0 to 65535: sam FLAG will be bitwise AND'd with this value, i.e. FLAG=FLAG & outSAMflagOR. This is applied after all flags have been set by STAR, but before outSAMflagOR. Can be used to unset specific bits that are not set otherwise.
--outSAMattrRGline	string multiple	SAM/BAM read group line. The first word contains the read group identifier and must start with "ID:", e.g. --outSAMattrRGline ID:xxx CN:yy "DS:z z z". xxx will be added as RG tag to each output alignment. Any spaces in the tag values have to be double quoted. Comma separated RG lines correspons to different (comma separated) input files in --readFilesIn. Commas have to be surrounded by spaces, e.g. --outSAMattrRGline ID:xxx , ID:zzz "DS:z z" , ID:yyy DS:yyyy
--outSAMheaderHD	string multiple	@HD (header) line of the SAM header
--outSAMheaderPG	string multiple	extra @PG (software) line of the SAM header (in addition to STAR)
--outSAMheaderCommentFile	string	path to the file with @CO (comment) lines of the SAM header
--outSAMfilter	string multiple	filter the output into main SAM/BAM files - KeepOnlyAddedReferences ... only keep the reads for which all alignments are to the extra reference sequences added with --genomeFastaFiles at the mapping stage. - KeepAllAddedReferences ... keep all alignments to the extra reference sequences added with --genomeFastaFiles at the mapping stage.
--outSAMmultNmax	integer	max number of multiple alignments for a read that will be output to the SAM/BAM files. Note that if this value is not equal to -1, the top scoring alignment will be output first - -1 ... all alignments (up to --outFilterMultimapNmax) will be output
--outSAMtlen	integer	calculation method for the TLEN field in the SAM/BAM files - 1 ... leftmost base of the (+)strand mate to rightmost base of the (-)mate. (+)sign for the (+)strand mate - 2 ... leftmost base of any mate to rightmost base of any mate. (+)sign for the mate with the leftmost base. This is different from 1 for overlapping mates with protruding ends
--outBAMcompression	integer	-1 to 10 BAM compression level, -1=default compression (6?), 0=no compression, 10=maximum compression
--outBAMsortingThreadN	integer	>=0: number of threads for BAM sorting. 0 will default to min(6,--runThreadN).
--outBAMsortingBinsN	integer	>0: number of genome bins for coordinate-sorting

Name

Type & Properties

--outSAMtype

string

multiple

--outSAMmode

string

--outSAMstrandField

string

--outSAMattributes

string

multiple

a string of desired SAM attributes, in the order desired for the output SAM. Tags can be listed in any combination/order. ***Presets: - None ... no attributes - Standard ... NH HI AS nM - All ... NH HI AS nM NM MD jM jI MC ch ***Alignment: - NH ... number of loci the reads maps to: =1 for unique mappers, >1 for multimappers. Standard SAM tag. - HI ... multiple alignment index, starts with --outSAMattrIHstart (=1 by default). Standard SAM tag. - AS ... local alignment score, +1/-1 for matches/mismateches, score* penalties for indels and gaps. For PE reads, total score for two mates. Stadnard SAM tag. - nM ... number of mismatches. For PE reads, sum over two mates. - NM ... edit distance to the reference (number of mismatched + inserted + deleted bases) for each mate. Standard SAM tag. - MD ... string encoding mismatched and deleted reference bases (see standard SAM specifications). Standard SAM tag. - jM ... intron motifs for all junctions (i.e. N in CIGAR): 0: non-canonical; 1: GT/AG, 2: CT/AC, 3: GC/AG, 4: CT/GC, 5: AT/AC, 6: GT/AT. If splice junctions database is used, and a junction is annotated, 20 is added to its motif value. - jI ... start and end of introns for all junctions (1-based). - XS ... alignment strand according to --outSAMstrandField. - MC ... mate's CIGAR string. Standard SAM tag. - ch ... marks all segment of all chimeric alingments for --chimOutType WithinBAM output. - cN ... number of bases clipped from the read ends: 5' and 3' ***Variation: - vA ... variant allele - vG ... genomic coordinate of the variant overlapped by the read. - vW ... 1 - alignment passes WASP filtering; 2,3,4,5,6,7 - alignment does not pass WASP filtering. Requires --waspOutputMode SAMtag. ***STARsolo: - CR CY UR UY ... sequences and quality scores of cell barcodes and UMIs for the solo* demultiplexing. - GX GN ... gene ID and gene name for unique-gene reads. - gx gn ... gene IDs and gene names for unique- and multi-gene reads. - CB UB ... error-corrected cell barcodes and UMIs for solo* demultiplexing. Requires --outSAMtype BAM SortedByCoordinate. - sM ... assessment of CB and UMI. - sS ... sequence of the entire barcode (CB,UMI,adapter). - sQ ... quality of the entire barcode. ***Unsupported/undocumented: - ha ... haplotype (1/2) when mapping to the diploid genome. Requires genome generated with --genomeTransformType Diploid . - rB ... alignment block read/genomic coordinates. - vR ... read coordinate of the variant.

--outSAMattrIHstart

integer

--outSAMunmapped

string

multiple

--outSAMorder

string

--outSAMprimaryFlag

string

--outSAMreadID

string

--outSAMmapqUnique

integer

--outSAMflagOR

integer

--outSAMflagAND

integer

--outSAMattrRGline

string

multiple

--outSAMheaderHD

string

multiple

--outSAMheaderPG

string

multiple

--outSAMheaderCommentFile

string

--outSAMfilter

string

multiple

--outSAMmultNmax

integer

--outSAMtlen

integer

--outBAMcompression

integer

--outBAMsortingThreadN

integer

--outBAMsortingBinsN

integer

BAM processing

Name	Type & Properties	Description
--bamRemoveDuplicatesType	string	mark duplicates in the BAM file, for now only works with (i) sorted BAM fed with inputBAMfile, and (ii) for paired-end alignments only - - ... no duplicate removal/marking - UniqueIdentical ... mark all multimappers, and duplicate unique mappers. The coordinates, FLAG, CIGAR must be identical - UniqueIdenticalNotMulti ... mark duplicate unique mappers but not multimappers.
--bamRemoveDuplicatesMate2basesN	integer	number of bases from the 5' of mate 2 to use in collapsing (e.g. for RAMPAGE)

Name

Type & Properties

--bamRemoveDuplicatesType

string

--bamRemoveDuplicatesMate2basesN

integer

Output Wiggle

Name	Type & Properties	Description
--outWigType	string multiple	type of signal output, e.g. "bedGraph" OR "bedGraph read1_5p". Requires sorted BAM: --outSAMtype BAM SortedByCoordinate . 1st word: - None ... no signal output - bedGraph ... bedGraph format - wiggle ... wiggle format 2nd word: - read1_5p ... signal from only 5' of the 1st read, useful for CAGE/RAMPAGE etc - read2 ... signal from only 2nd read
--outWigStrand	string	strandedness of wiggle/bedGraph output - Stranded ... separate strands, str1 and str2 - Unstranded ... collapsed strands
--outWigReferencesPrefix	string	prefix matching reference names to include in the output wiggle file, e.g. "chr", default "-" - include all references
--outWigNorm	string	type of normalization for the signal - RPM ... reads per million of mapped reads - None ... no normalization, "raw" counts

Name

Type & Properties

--outWigType

string

multiple

--outWigStrand

string

--outWigReferencesPrefix

string

--outWigNorm

string

Output Filtering

Name	Type & Properties	Description
--outFilterType	string	type of filtering - Normal ... standard filtering using only current alignment - BySJout ... keep only those reads that contain junctions that passed filtering into SJ.out.tab
--outFilterMultimapScoreRange	integer	the score range below the maximum score for multimapping alignments
--outFilterMultimapNmax	integer	maximum number of loci the read is allowed to map to. Alignments (all of them) will be output only if the read maps to no more loci than this value. Otherwise no alignments will be output, and the read will be counted as "mapped to too many loci" in the Log.final.out .
--outFilterMismatchNmax	integer	alignment will be output only if it has no more mismatches than this value.
--outFilterMismatchNoverLmax	double	alignment will be output only if its ratio of mismatches to mapped length is less than or equal to this value.
--outFilterMismatchNoverReadLmax	double	alignment will be output only if its ratio of mismatches to read length is less than or equal to this value.
--outFilterScoreMin	integer	alignment will be output only if its score is higher than or equal to this value.
--outFilterScoreMinOverLread	double	same as outFilterScoreMin, but normalized to read length (sum of mates' lengths for paired-end reads)
--outFilterMatchNmin	integer	alignment will be output only if the number of matched bases is higher than or equal to this value.
--outFilterMatchNminOverLread	double	sam as outFilterMatchNmin, but normalized to the read length (sum of mates' lengths for paired-end reads).
--outFilterIntronMotifs	string	filter alignment using their motifs - None ... no filtering - RemoveNoncanonical ... filter out alignments that contain non-canonical junctions - RemoveNoncanonicalUnannotated ... filter out alignments that contain non-canonical unannotated junctions when using annotated splice junctions database. The annotated non-canonical junctions will be kept.
--outFilterIntronStrands	string	filter alignments - RemoveInconsistentStrands ... remove alignments that have junctions with inconsistent strands - None ... no filtering

Name

Type & Properties

--outFilterType

string

--outFilterMultimapScoreRange

integer

--outFilterMultimapNmax

integer

--outFilterMismatchNmax

integer

--outFilterMismatchNoverLmax

double

--outFilterMismatchNoverReadLmax

double

--outFilterScoreMin

integer

--outFilterScoreMinOverLread

double

--outFilterMatchNmin

integer

--outFilterMatchNminOverLread

double

--outFilterIntronMotifs

string

--outFilterIntronStrands

string

Output splice junctions (SJ.out.tab)

Name	Type & Properties	Description
--outSJtype	string	type of splice junction output - Standard ... standard SJ.out.tab output - None ... no splice junction output

Name

Type & Properties

--outSJtype

string

Output Filtering: Splice Junctions

Name	Type & Properties	Description
--outSJfilterReads	string	which reads to consider for collapsed splice junctions output - All ... all reads, unique- and multi-mappers - Unique ... uniquely mapping reads only
--outSJfilterOverhangMin	integer multiple	minimum overhang length for splice junctions on both sides for: (1) non-canonical motifs, (2) GT/AG and CT/AC motif, (3) GC/AG and CT/GC motif, (4) AT/AC and GT/AT motif. -1 means no output for that motif does not apply to annotated junctions
--outSJfilterCountUniqueMin	integer multiple	minimum uniquely mapping read count per junction for: (1) non-canonical motifs, (2) GT/AG and CT/AC motif, (3) GC/AG and CT/GC motif, (4) AT/AC and GT/AT motif. -1 means no output for that motif Junctions are output if one of outSJfilterCountUniqueMin OR outSJfilterCountTotalMin conditions are satisfied does not apply to annotated junctions
--outSJfilterCountTotalMin	integer multiple	minimum total (multi-mapping+unique) read count per junction for: (1) non-canonical motifs, (2) GT/AG and CT/AC motif, (3) GC/AG and CT/GC motif, (4) AT/AC and GT/AT motif. -1 means no output for that motif Junctions are output if one of outSJfilterCountUniqueMin OR outSJfilterCountTotalMin conditions are satisfied does not apply to annotated junctions
--outSJfilterDistToOtherSJmin	integer multiple	minimum allowed distance to other junctions' donor/acceptor does not apply to annotated junctions
--outSJfilterIntronMaxVsReadN	integer multiple	maximum gap allowed for junctions supported by 1,2,3,,,N reads i.e. by default junctions supported by 1 read can have gaps <=50000b, by 2 reads: <=100000b, by 3 reads: <=200000. by >=4 reads any gap <=alignIntronMax does not apply to annotated junctions

Name

Type & Properties

--outSJfilterReads

string

--outSJfilterOverhangMin

integer

multiple

--outSJfilterCountUniqueMin

integer

multiple

--outSJfilterCountTotalMin

integer

multiple

--outSJfilterDistToOtherSJmin

integer

multiple

--outSJfilterIntronMaxVsReadN

integer

multiple

Scoring

Name	Type & Properties	Description
--scoreGap	integer	splice junction penalty (independent on intron motif)
--scoreGapNoncan	integer	non-canonical junction penalty (in addition to scoreGap)
--scoreGapGCAG	integer	GC/AG and CT/GC junction penalty (in addition to scoreGap)
--scoreGapATAC	integer	AT/AC and GT/AT junction penalty (in addition to scoreGap)
--scoreGenomicLengthLog2scale	integer	extra score logarithmically scaled with genomic length of the alignment: scoreGenomicLengthLog2scale*log2(genomicLength)
--scoreDelOpen	integer	deletion open penalty
--scoreDelBase	integer	deletion extension penalty per base (in addition to scoreDelOpen)
--scoreInsOpen	integer	insertion open penalty
--scoreInsBase	integer	insertion extension penalty per base (in addition to scoreInsOpen)
--scoreStitchSJshift	integer	maximum score reduction while searching for SJ boundaries in the stitching step

Name

Type & Properties

--scoreGap

integer

--scoreGapNoncan

integer

--scoreGapGCAG

integer

--scoreGapATAC

integer

--scoreGenomicLengthLog2scale

integer

--scoreDelOpen

integer

--scoreDelBase

integer

--scoreInsOpen

integer

--scoreInsBase

integer

--scoreStitchSJshift

integer

Alignments and Seeding

Name	Type & Properties	Description
--seedSearchStartLmax	integer	defines the search start point through the read - the read is split into pieces no longer than this value
--seedSearchStartLmaxOverLread	double	seedSearchStartLmax normalized to read length (sum of mates' lengths for paired-end reads)
--seedSearchLmax	integer	defines the maximum length of the seeds, if =0 seed length is not limited
--seedMultimapNmax	integer	only pieces that map fewer than this value are utilized in the stitching procedure
--seedPerReadNmax	integer	max number of seeds per read
--seedPerWindowNmax	integer	max number of seeds per window
--seedNoneLociPerWindow	integer	max number of one seed loci per window
--seedSplitMin	integer	min length of the seed sequences split by Ns or mate gap
--seedMapMin	integer	min length of seeds to be mapped
--alignIntronMin	integer	minimum intron size, genomic gap is considered intron if its length>=alignIntronMin, otherwise it is considered Deletion
--alignIntronMax	integer	maximum intron size, if 0, max intron size will be determined by (2^winBinNbits)*winAnchorDistNbins
--alignMatesGapMax	integer	maximum gap between two mates, if 0, max intron gap will be determined by (2^winBinNbits)*winAnchorDistNbins
--alignSJoverhangMin	integer	minimum overhang (i.e. block size) for spliced alignments
--alignSJstitchMismatchNmax	integer multiple	maximum number of mismatches for stitching of the splice junctions (-1: no limit). (1) non-canonical motifs, (2) GT/AG and CT/AC motif, (3) GC/AG and CT/GC motif, (4) AT/AC and GT/AT motif.
--alignSJDBoverhangMin	integer	minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments
--alignSplicedMateMapLmin	integer	minimum mapped length for a read mate that is spliced
--alignSplicedMateMapLminOverLmate	double	alignSplicedMateMapLmin normalized to mate length
--alignWindowsPerReadNmax	integer	max number of windows per read
--alignTranscriptsPerWindowNmax	integer	max number of transcripts per window
--alignTranscriptsPerReadNmax	integer	max number of different alignments per read to consider
--alignEndsType	string	type of read ends alignment - Local ... standard local alignment with soft-clipping allowed - EndToEnd ... force end-to-end read alignment, do not soft-clip - Extend5pOfRead1 ... fully extend only the 5p of the read1, all other ends: local alignment - Extend5pOfReads12 ... fully extend only the 5p of the both read1 and read2, all other ends: local alignment
--alignEndsProtrude	string	allow protrusion of alignment ends, i.e. start (end) of the +strand mate downstream of the start (end) of the -strand mate 1st word: int: maximum number of protrusion bases allowed 2nd word: string: - ConcordantPair ... report alignments with non-zero protrusion as concordant pairs - DiscordantPair ... report alignments with non-zero protrusion as discordant pairs
--alignSoftClipAtReferenceEnds	string	allow the soft-clipping of the alignments past the end of the chromosomes - Yes ... allow - No ... prohibit, useful for compatibility with Cufflinks
--alignInsertionFlush	string	how to flush ambiguous insertion positions - None ... insertions are not flushed - Right ... insertions are flushed to the right

Name

Type & Properties

--seedSearchStartLmax

integer

--seedSearchStartLmaxOverLread

double

--seedSearchLmax

integer

--seedMultimapNmax

integer

--seedPerReadNmax

integer

--seedPerWindowNmax

integer

--seedNoneLociPerWindow

integer

--seedSplitMin

integer

--seedMapMin

integer

--alignIntronMin

integer

--alignIntronMax

integer

--alignMatesGapMax

integer

--alignSJoverhangMin

integer

--alignSJstitchMismatchNmax

integer

multiple

--alignSJDBoverhangMin

integer

--alignSplicedMateMapLmin

integer

--alignSplicedMateMapLminOverLmate

double

--alignWindowsPerReadNmax

integer

--alignTranscriptsPerWindowNmax

integer

--alignTranscriptsPerReadNmax

integer

--alignEndsType

string

--alignEndsProtrude

string

--alignSoftClipAtReferenceEnds

string

--alignInsertionFlush

string

Paired-End reads

Name	Type & Properties	Description
--peOverlapNbasesMin	integer	minimum number of overlapping bases to trigger mates merging and realignment. Specify >0 value to switch on the "merginf of overlapping mates" algorithm.
--peOverlapMMp	double	maximum proportion of mismatched bases in the overlap area

Name

Type & Properties

--peOverlapNbasesMin

integer

--peOverlapMMp

double

Windows, Anchors, Binning

Name	Type & Properties	Description
--winAnchorMultimapNmax	integer	max number of loci anchors are allowed to map to
--winBinNbits	integer	=log2(winBin), where winBin is the size of the bin for the windows/clustering, each window will occupy an integer number of bins.
--winAnchorDistNbins	integer	max number of bins between two anchors that allows aggregation of anchors into one window
--winFlankNbins	integer	log2(winFlank), where win Flank is the size of the left and right flanking regions for each window
--winReadCoverageRelativeMin	double	minimum relative coverage of the read sequence by the seeds in a window, for STARlong algorithm only.
--winReadCoverageBasesMin	integer	minimum number of bases covered by the seeds in a window , for STARlong algorithm only.

Name

Type & Properties

--winAnchorMultimapNmax

integer

--winBinNbits

integer

--winAnchorDistNbins

integer

--winFlankNbins

integer

--winReadCoverageRelativeMin

double

--winReadCoverageBasesMin

integer

Chimeric Alignments

Name	Type & Properties	Description
--chimOutType	string multiple	type of chimeric output - Junctions ... Chimeric.out.junction - SeparateSAMold ... output old SAM into separate Chimeric.out.sam file - WithinBAM ... output into main aligned BAM files (Aligned.*.bam) - WithinBAM HardClip ... (default) hard-clipping in the CIGAR for supplemental chimeric alignments (default if no 2nd word is present) - WithinBAM SoftClip ... soft-clipping in the CIGAR for supplemental chimeric alignments
--chimSegmentMin	integer	minimum length of chimeric segment length, if ==0, no chimeric output
--chimScoreMin	integer	minimum total (summed) score of the chimeric segments
--chimScoreDropMax	integer	max drop (difference) of chimeric score (the sum of scores of all chimeric segments) from the read length
--chimScoreSeparation	integer	minimum difference (separation) between the best chimeric score and the next one
--chimScoreJunctionNonGTAG	integer	penalty for a non-GT/AG chimeric junction
--chimJunctionOverhangMin	integer	minimum overhang for a chimeric junction
--chimSegmentReadGapMax	integer	maximum gap in the read sequence between chimeric segments
--chimFilter	string multiple	different filters for chimeric alignments - None ... no filtering - banGenomicN ... Ns are not allowed in the genome sequence around the chimeric junction
--chimMainSegmentMultNmax	integer	maximum number of multi-alignments for the main chimeric segment. =1 will prohibit multimapping main segments.
--chimMultimapNmax	integer	maximum number of chimeric multi-alignments - 0 ... use the old scheme for chimeric detection which only considered unique alignments
--chimMultimapScoreRange	integer	the score range for multi-mapping chimeras below the best chimeric score. Only works with --chimMultimapNmax > 1
--chimNonchimScoreDropMin	integer	to trigger chimeric detection, the drop in the best non-chimeric alignment score with respect to the read length has to be greater than this value
--chimOutJunctionFormat	integer	formatting type for the Chimeric.out.junction file - 0 ... no comment lines/headers - 1 ... comment lines at the end of the file: command line and Nreads: total, unique/multi-mapping

Name

Type & Properties

--chimOutType

string

multiple

--chimSegmentMin

integer

--chimScoreMin

integer

--chimScoreDropMax

integer

--chimScoreSeparation

integer

--chimScoreJunctionNonGTAG

integer

--chimJunctionOverhangMin

integer

--chimSegmentReadGapMax

integer

--chimFilter

string

multiple

--chimMainSegmentMultNmax

integer

--chimMultimapNmax

integer

--chimMultimapScoreRange

integer

--chimNonchimScoreDropMin

integer

--chimOutJunctionFormat

integer

Quantification of Annotations

Name	Type & Properties	Description
--quantMode	string multiple	types of quantification requested - - ... none - TranscriptomeSAM ... output SAM/BAM alignments to transcriptome into a separate file - GeneCounts ... count reads per gene
--quantTranscriptomeBAMcompression	integer	-2 to 10 transcriptome BAM compression level - -2 ... no BAM output - -1 ... default compression (6?) - 0 ... no compression - 10 ... maximum compression
--quantTranscriptomeBan	string	prohibit various alignment type - IndelSoftclipSingleend ... prohibit indels, soft clipping and single-end alignments - compatible with RSEM - Singleend ... prohibit single-end alignments

Name

Type & Properties

--quantMode

string

multiple

--quantTranscriptomeBAMcompression

integer

--quantTranscriptomeBan

string

2-pass Mapping

Name	Type & Properties	Description
--twopassMode	string	2-pass mapping mode. - None ... 1-pass mapping - Basic ... basic 2-pass mapping, with all 1st pass junctions inserted into the genome indices on the fly
--twopass1readsN	integer	number of reads to process for the 1st step. Use very large number (or default -1) to map all reads in the first step.

Name

Type & Properties

--twopassMode

string

--twopass1readsN

integer

WASP parameters

Name	Type & Properties	Description
--waspOutputMode	string	WASP allele-specific output type. This is re-implementation of the original WASP mappability filtering by Bryce van de Geijn, Graham McVicker, Yoav Gilad & Jonathan K Pritchard. Please cite the original WASP paper: Nature Methods 12, 1061-1063 (2015), https://www.nature.com/articles/nmeth.3582 . - SAMtag ... add WASP tags to the alignments that pass WASP filtering

Name

Type & Properties

--waspOutputMode

string

STARsolo (single cell RNA-seq) parameters

Name	Type & Properties	Description
--soloType	string multiple	type of single-cell RNA-seq - CB_UMI_Simple ... (a.k.a. Droplet) one UMI and one Cell Barcode of fixed length in read2, e.g. Drop-seq and 10X Chromium. - CB_UMI_Complex ... multiple Cell Barcodes of varying length, one UMI of fixed length and one adapter sequence of fixed length are allowed in read2 only (e.g. inDrop, ddSeq). - CB_samTagOut ... output Cell Barcode as CR and/or CB SAm tag. No UMI counting. --readFilesIn cDNA_read1 [cDNA_read2 if paired-end] CellBarcode_read . Requires --outSAMtype BAM Unsorted [and/or SortedByCoordinate] - SmartSeq ... Smart-seq: each cell in a separate FASTQ (paired- or single-end), barcodes are corresponding read-groups, no UMI sequences, alignments deduplicated according to alignment start and end (after extending soft-clipped bases)
--soloCBwhitelist	string multiple	file(s) with whitelist(s) of cell barcodes. Only --soloType CB_UMI_Complex allows more than one whitelist file. - None ... no whitelist: all cell barcodes are allowed
--soloCBstart	integer	cell barcode start base
--soloCBlen	integer	cell barcode length
--soloUMIstart	integer	UMI start base
--soloUMIlen	integer	UMI length
--soloBarcodeReadLength	integer	length of the barcode read - 1 ... equal to sum of soloCBlen+soloUMIlen - 0 ... not defined, do not check
--soloBarcodeMate	integer	identifies which read mate contains the barcode (CB+UMI) sequence - 0 ... barcode sequence is on separate read, which should always be the last file in the --readFilesIn listed - 1 ... barcode sequence is a part of mate 1 - 2 ... barcode sequence is a part of mate 2
--soloCBposition	string multiple	position of Cell Barcode(s) on the barcode read. Presently only works with --soloType CB_UMI_Complex, and barcodes are assumed to be on Read2. Format for each barcode: startAnchor_startPosition_endAnchor_endPosition start(end)Anchor defines the Anchor Base for the CB: 0: read start; 1: read end; 2: adapter start; 3: adapter end start(end)Position is the 0-based position with of the CB start(end) with respect to the Anchor Base String for different barcodes are separated by space. Example: inDrop (Zilionis et al, Nat. Protocols, 2017): --soloCBposition 0_0_2_-1 3_1_3_8
--soloUMIposition	string	position of the UMI on the barcode read, same as soloCBposition Example: inDrop (Zilionis et al, Nat. Protocols, 2017): --soloCBposition 3_9_3_14
--soloAdapterSequence	string	adapter sequence to anchor barcodes. Only one adapter sequence is allowed.
--soloAdapterMismatchesNmax	integer	maximum number of mismatches allowed in adapter sequence.
--soloCBmatchWLtype	string	matching the Cell Barcodes to the WhiteList - Exact ... only exact matches allowed - 1MM ... only one match in whitelist with 1 mismatched base allowed. Allowed CBs have to have at least one read with exact match. - 1MM_multi ... multiple matches in whitelist with 1 mismatched base allowed, posterior probability calculation is used choose one of the matches. Allowed CBs have to have at least one read with exact match. This option matches best with CellRanger 2.2.0 - 1MM_multi_pseudocounts ... same as 1MM_Multi, but pseudocounts of 1 are added to all whitelist barcodes. - 1MM_multi_Nbase_pseudocounts ... same as 1MM_multi_pseudocounts, multimatching to WL is allowed for CBs with N-bases. This option matches best with CellRanger >= 3.0.0 - EditDist_2 ... allow up to edit distance of 3 fpr each of the barcodes. May include one deletion + one insertion. Only works with --soloType CB_UMI_Complex. Matches to multiple passlist barcdoes are not allowed. Similar to ParseBio Split-seq pipeline.
--soloInputSAMattrBarcodeSeq	string multiple	when inputting reads from a SAM file (--readsFileType SAM SE/PE), these SAM attributes mark the barcode sequence (in proper order). For instance, for 10X CellRanger or STARsolo BAMs, use --soloInputSAMattrBarcodeSeq CR UR . This parameter is required when running STARsolo with input from SAM.
--soloInputSAMattrBarcodeQual	string multiple	when inputting reads from a SAM file (--readsFileType SAM SE/PE), these SAM attributes mark the barcode qualities (in proper order). For instance, for 10X CellRanger or STARsolo BAMs, use --soloInputSAMattrBarcodeQual CY UY . If this parameter is '-' (default), the quality 'H' will be assigned to all bases.
--soloStrand	string	strandedness of the solo libraries: - Unstranded ... no strand information - Forward ... read strand same as the original RNA molecule - Reverse ... read strand opposite to the original RNA molecule
--soloFeatures	string multiple	genomic features for which the UMI counts per Cell Barcode are collected - Gene ... genes: reads match the gene transcript - SJ ... splice junctions: reported in SJ.out.tab - GeneFull ... full gene (pre-mRNA): count all reads overlapping genes' exons and introns - GeneFull_ExonOverIntron ... full gene (pre-mRNA): count all reads overlapping genes' exons and introns: prioritize 100% overlap with exons - GeneFull_Ex50pAS ... full gene (pre-RNA): count all reads overlapping genes' exons and introns: prioritize >50% overlap with exons. Do not count reads with 100% exonic overlap in the antisense direction.
--soloMultiMappers	string multiple	counting method for reads mapping to multiple genes - Unique ... count only reads that map to unique genes - Uniform ... uniformly distribute multi-genic UMIs to all genes - Rescue ... distribute UMIs proportionally to unique+uniform counts (~ first iteration of EM) - PropUnique ... distribute UMIs proportionally to unique mappers, if present, and uniformly if not. - EM ... multi-gene UMIs are distributed using Expectation Maximization algorithm
--soloUMIdedup	string multiple	type of UMI deduplication (collapsing) algorithm - 1MM_All ... all UMIs with 1 mismatch distance to each other are collapsed (i.e. counted once). - 1MM_Directional_UMItools ... follows the "directional" method from the UMI-tools by Smith, Heger and Sudbery (Genome Research 2017). - 1MM_Directional ... same as 1MM_Directional_UMItools, but with more stringent criteria for duplicate UMIs - Exact ... only exactly matching UMIs are collapsed. - NoDedup ... no deduplication of UMIs, count all reads. - 1MM_CR ... CellRanger2-4 algorithm for 1MM UMI collapsing.
--soloUMIfiltering	string multiple	type of UMI filtering (for reads uniquely mapping to genes) - - ... basic filtering: remove UMIs with N and homopolymers (similar to CellRanger 2.2.0). - MultiGeneUMI ... basic + remove lower-count UMIs that map to more than one gene. - MultiGeneUMI_All ... basic + remove all UMIs that map to more than one gene. - MultiGeneUMI_CR ... basic + remove lower-count UMIs that map to more than one gene, matching CellRanger > 3.0.0 . Only works with --soloUMIdedup 1MM_CR
--soloOutFileNames	string multiple	file names for STARsolo output: file_name_prefix gene_names barcode_sequences cell_feature_count_matrix
--soloCellFilter	string multiple	cell filtering type and parameters - None ... do not output filtered cells - TopCells ... only report top cells by UMI count, followed by the exact number of cells - CellRanger2.2 ... simple filtering of CellRanger 2.2. Can be followed by numbers: number of expected cells, robust maximum percentile for UMI count, maximum to minimum ratio for UMI count The harcoded values are from CellRanger: nExpectedCells=3000; maxPercentile=0.99; maxMinRatio=10 - EmptyDrops_CR ... EmptyDrops filtering in CellRanger flavor. Please cite the original EmptyDrops paper: A.T.L Lun et al, Genome Biology, 20, 63 (2019): https://genomebiology.biomedcentral.com/articles/10.1186/s13059-019-1662-y Can be followed by 10 numeric parameters: nExpectedCells maxPercentile maxMinRatio indMin indMax umiMin umiMinFracMedian candMaxN FDR simN The harcoded values are from CellRanger: 3000 0.99 10 45000 90000 500 0.01 20000 0.01 10000
--soloOutFormatFeaturesGeneField3	string multiple	field 3 in the Gene features.tsv file. If "-", then no 3rd field is output.
--soloCellReadStats	string	Output reads statistics for each CB - Standard ... standard output

Name

Type & Properties

--soloType

string

multiple

--soloCBwhitelist

string

multiple

--soloCBstart

integer

--soloCBlen

integer

--soloUMIstart

integer

--soloUMIlen

integer

--soloBarcodeReadLength

integer

--soloBarcodeMate

integer

--soloCBposition

string

multiple

--soloUMIposition

string

--soloAdapterSequence

string

--soloAdapterMismatchesNmax

integer

--soloCBmatchWLtype

string

--soloInputSAMattrBarcodeSeq

string

multiple

--soloInputSAMattrBarcodeQual

string

multiple

--soloStrand

string

--soloFeatures

string

multiple

--soloMultiMappers

string

multiple

--soloUMIdedup

string

multiple

--soloUMIfiltering

string

multiple

--soloOutFileNames

string

multiple

--soloCellFilter

string

multiple

--soloOutFormatFeaturesGeneField3

string

multiple

--soloCellReadStats

string