Program options for bowtie2_build: Bowtie 2 version 2.4.2 by Ben Langmead (langmea@cs.jhu.edu, www.cs.jhu.edu/~langmea) Usage: bowtie2-build [options]* reference_in comma-separated list of files with ref sequences bt2_index_base write bt2 data to files with this dir/basename *** Bowtie 2 indexes work only with v2 (not v1). Likewise for v1 indexes. *** Options: -f reference files are Fasta (default) -c reference sequences given on cmd line (as ) --large-index force generated index to be 'large', even if ref has fewer than 4 billion nucleotides --debug use the debug binary; slower, assertions enabled --sanitized use sanitized binary; slower, uses ASan and/or UBSan --verbose log the issued command -a/--noauto disable automatic -p/--bmax/--dcv memory-fitting -p/--packed use packed strings internally; slower, less memory --bmax max bucket sz for blockwise suffix-array builder --bmaxdivn max bucket sz as divisor of ref len (default: 4) --dcv diff-cover period for blockwise (default: 1024) --nodc disable diff-cover (algorithm becomes quadratic) -r/--noref don't build .3/.4 index files -3/--justref just build .3/.4 index files -o/--offrate SA is sampled every 2^ BWT chars (default: 5) -t/--ftabchars # of chars consumed in initial lookup (default: 10) --threads # of threads --seed seed for random number generator -q/--quiet verbose output (for debugging) -h/--help print detailed description of tool and its options --usage print this usage message --version print version information and quit ---------------------------------------------- Program options for bowtie2: Bowtie 2 version 2.4.2 by Ben Langmead (langmea@cs.jhu.edu, www.cs.jhu.edu/~langmea) Usage: bowtie2 [options]* -x {-1 -2 | -U | --interleaved | -b } [-S ] Index filename prefix (minus trailing .X.bt2). NOTE: Bowtie 1 and Bowtie 2 indexes are not compatible. Files with #1 mates, paired with files in . Could be gzip'ed (extension: .gz) or bzip2'ed (extension: .bz2). Files with #2 mates, paired with files in . Could be gzip'ed (extension: .gz) or bzip2'ed (extension: .bz2). Files with unpaired reads. Could be gzip'ed (extension: .gz) or bzip2'ed (extension: .bz2). Files with interleaved paired-end FASTQ/FASTA reads Could be gzip'ed (extension: .gz) or bzip2'ed (extension: .bz2). Files are unaligned BAM sorted by read name. File for SAM output (default: stdout) , , can be comma-separated lists (no whitespace) and can be specified many times. E.g. '-U file1.fq,file2.fq -U file3.fq'. Options (defaults in parentheses): Input: -q query input files are FASTQ .fq/.fastq (default) --tab5 query input files are TAB5 .tab5 --tab6 query input files are TAB6 .tab6 --qseq query input files are in Illumina's qseq format -f query input files are (multi-)FASTA .fa/.mfa -r query input files are raw one-sequence-per-line -F k:,i: query input files are continuous FASTA where reads are substrings (k-mers) extracted from a FASTA file and aligned at offsets 1, 1+i, 1+2i ... end of reference -c , , are sequences themselves, not files -s/--skip skip the first reads/pairs in the input (none) -u/--upto stop after first reads/pairs (no limit) -5/--trim5 trim bases from 5'/left end of reads (0) -3/--trim3 trim bases from 3'/right end of reads (0) --trim-to [3:|5:] trim reads exceeding bases from either 3' or 5' end If the read end is not specified then it defaults to 3 (0) --phred33 qualities are Phred+33 (default) --phred64 qualities are Phred+64 --int-quals qualities encoded as space-delimited integers Presets: Same as: For --end-to-end: --very-fast -D 5 -R 1 -N 0 -L 22 -i S,0,2.50 --fast -D 10 -R 2 -N 0 -L 22 -i S,0,2.50 --sensitive -D 15 -R 2 -N 0 -L 22 -i S,1,1.15 (default) --very-sensitive -D 20 -R 3 -N 0 -L 20 -i S,1,0.50 For --local: --very-fast-local -D 5 -R 1 -N 0 -L 25 -i S,1,2.00 --fast-local -D 10 -R 2 -N 0 -L 22 -i S,1,1.75 --sensitive-local -D 15 -R 2 -N 0 -L 20 -i S,1,0.75 (default) --very-sensitive-local -D 20 -R 3 -N 0 -L 20 -i S,1,0.50 Alignment: -N max # mismatches in seed alignment; can be 0 or 1 (0) -L length of seed substrings; must be >3, <32 (22) -i interval between seed substrings w/r/t read len (S,1,1.15) --n-ceil func for max # non-A/C/G/Ts permitted in aln (L,0,0.15) --dpad include extra ref chars on sides of DP table (15) --gbar disallow gaps within nucs of read extremes (4) --ignore-quals treat all quality values as 30 on Phred scale (off) --nofw do not align forward (original) version of read (off) --norc do not align reverse-complement version of read (off) --no-1mm-upfront do not allow 1 mismatch alignments before attempting to scan for the optimal seeded alignments --end-to-end entire read must align; no clipping (on) OR --local local alignment; ends might be soft clipped (off) Scoring: --ma match bonus (0 for --end-to-end, 2 for --local) --mp max penalty for mismatch; lower qual = lower penalty (6) --np penalty for non-A/C/G/Ts in read/ref (1) --rdg , read gap open, extend penalties (5,3) --rfg , reference gap open, extend penalties (5,3) --score-min min acceptable alignment score w/r/t read length (G,20,8 for local, L,-0.6,-0.6 for end-to-end) Reporting: (default) look for multiple alignments, report best, with MAPQ OR -k report up to alns per read; MAPQ not meaningful OR -a/--all report all alignments; very slow, MAPQ not meaningful Effort: -D give up extending after failed extends in a row (15) -R for reads w/ repetitive seeds, try sets of seeds (2) Paired-end: -I/--minins minimum fragment length (0) -X/--maxins maximum fragment length (500) --fr/--rf/--ff -1, -2 mates align fw/rev, rev/fw, fw/fw (--fr) --no-mixed suppress unpaired alignments for paired reads --no-discordant suppress discordant alignments for paired reads --dovetail concordant when mates extend past each other --no-contain not concordant when one mate alignment contains other --no-overlap not concordant when mates overlap at all BAM: --align-paired-reads Bowtie2 will, by default, attempt to align unpaired BAM reads. Use this option to align paired-end reads instead. --preserve-tags Preserve tags from the original BAM record by appending them to the end of the corresponding SAM output. Output: -t/--time print wall-clock time taken by search phases --un write unpaired reads that didn't align to --al write unpaired reads that aligned at least once to --un-conc write pairs that didn't align concordantly to --al-conc write pairs that aligned concordantly at least once to (Note: for --un, --al, --un-conc, or --al-conc, add '-gz' to the option name, e.g. --un-gz , to gzip compress output, or add '-bz2' to bzip2 compress output.) --quiet print nothing to stderr except serious errors --met-file send metrics to file at (off) --met-stderr send metrics to stderr (off) --met report internal counters & metrics every secs (1) --no-unal suppress SAM records for unaligned reads --no-head suppress header lines, i.e. lines starting with @ --no-sq suppress @SQ header lines --rg-id set read group id, reflected in @RG line and RG:Z: opt field --rg add ("lab:value") to @RG line of SAM header. Note: @RG line only printed when --rg-id is set. --omit-sec-seq put '*' in SEQ and QUAL fields for secondary alignments. --sam-no-qname-trunc Suppress standard behavior of truncating readname at first whitespace at the expense of generating non-standard SAM. --xeq Use '='/'X', instead of 'M,' to specify matches/mismatches in SAM record. --soft-clipped-unmapped-tlen Exclude soft-clipped bases when reporting TLEN --sam-append-comment Append FASTA/FASTQ comment to SAM record Performance: -p/--threads number of alignment threads to launch (1) --reorder force SAM output order to match order of input reads --mm use memory-mapped I/O for index; many 'bowtie's can share Other: --qc-filter filter out reads that are bad according to QSEQ filter --seed seed for random number generator (0) --non-deterministic seed rand. gen. arbitrarily instead of using read attributes --version print version information and quit -h/--help print this usage message ---------------------------------------------- Program options for samtools_sort: samtools sort: failed to read header from "-" sort: invalid option -- 'h' Usage: samtools sort [options...] [in.bam] Options: -l INT Set compression level, from 0 (uncompressed) to 9 (best) -m INT Set maximum memory per thread; suffix K/M/G recognized [768M] -n Sort by read name -t TAG Sort by value of TAG. Uses position as secondary index (or read name if -n is set) -o FILE Write final output to FILE rather than standard output -T PREFIX Write temporary files to PREFIX.nnnn.bam --no-PG do not add a PG line --input-fmt-option OPT[=VAL] Specify a single input file format option in the form of OPTION or OPTION=VALUE -O, --output-fmt FORMAT[,OPT[=VAL]]... Specify output format (SAM, BAM, CRAM) --output-fmt-option OPT[=VAL] Specify a single output file format option in the form of OPTION or OPTION=VALUE --reference FILE Reference sequence FASTA FILE [null] -@, --threads INT Number of additional threads to use [0] --verbosity INT Set level of verbosity ---------------------------------------------- Program options for samtools_view: [main_samview] fail to read the header from "-". [main_samview] fail to read the header from "-". ---------------------------------------------- Program options for samtools_index: Usage: samtools index [-bc] [-m INT] [out.index] Options: -b Generate BAI-format index for BAM files [default] -c Generate CSI-format index for BAM files -m INT Set minimum interval size for CSI indices to 2^INT [14] -@ INT Sets the number of threads [none] index: invalid option -- 'h' Usage: samtools index [-bc] [-m INT] [out.index] Options: -b Generate BAI-format index for BAM files [default] -c Generate CSI-format index for BAM files -m INT Set minimum interval size for CSI indices to 2^INT [14] -@ INT Sets the number of threads [none] ----------------------------------------------