hisat2-align-s - graph-based alignment of short nucleotide reads
to many genomes, wrapper script
HISAT2 version 2.2.1 by Daehwan Kim (infphilo@gmail.com,
www.ccb.jhu.edu/people/infphilo) Usage:
- hisat2 [options]* -x <ht2-idx> {-1 <m1> -2
<m2> | -U <r>} [-S <sam>]
- <ht2-idx>
- Index filename prefix (minus trailing .X.ht2).
- <m1>
- Files with #1 mates, paired with files in <m2>. Could be gzip'ed
(extension: .gz) or bzip2'ed (extension: .bz2).
- <m2>
- Files with #2 mates, paired with files in <m1>. Could be gzip'ed
(extension: .gz) or bzip2'ed (extension: .bz2).
- <r>
- Files with unpaired reads. Could be gzip'ed (extension: .gz) or bzip2'ed
(extension: .bz2).
- <sam>
- File for SAM output (default: stdout)
- <m1>, <m2>, <r> 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)
- --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
- -c
- <m1>, <m2>, <r> are sequences themselves, not files
- -s/--skip
<int>
- skip the first <int> reads/pairs in the input (none)
- -u/--upto
<int>
- stop after first <int> reads/pairs (no limit)
- -5/--trim5 <int>
- trim <int> bases from 5'/left end of reads (0)
- -3/--trim3 <int>
- trim <int> bases from 3'/right end of reads (0)
- --phred33
- qualities are Phred+33 (default)
- --phred64
- qualities are Phred+64
- --int-quals
- qualities encoded as space-delimited integers
- Presets:
- Same as:
--fast --no-repeat-index
--sensitive --bowtie2-dp 1 -k 30
--score-min L,0,-0.5
--very-sensitive --bowtie2-dp 2 -k 50
--score-min L,0,-1
- Alignment:
--bowtie2-dp <int> use Bowtie2's dynamic
programming alignment algorithm (0) - 0: no dynamic programming, 1:
conditional dynamic programming, and 2: unconditional dynamic programming
(slowest)
- --n-ceil
<func>
- func for max # non-A/C/G/Ts permitted in aln (L,0,0.15)
- --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-repeat-index
- do not use repeat index
- Spliced Alignment:
- --pen-cansplice
<int>
- penalty for a canonical splice site (0)
- --pen-noncansplice
<int>
- penalty for a non-canonical splice site (12)
- --pen-canintronlen
<func>
- penalty for long introns (G,-8,1) with canonical splice sites
- --pen-noncanintronlen
<func>
- penalty for long introns (G,-8,1) with noncanonical splice sites
- --min-intronlen
<int>
- minimum intron length (20)
- --max-intronlen
<int>
- maximum intron length (500000)
- --known-splicesite-infile
<path>
- provide a list of known splice sites
- --novel-splicesite-outfile
<path>
- report a list of splice sites
- --novel-splicesite-infile
<path>
- provide a list of novel splice sites
- --no-temp-splicesite
- disable the use of splice sites found
- --no-spliced-alignment
- disable spliced alignment
- --rna-strandness
<string>
- specify strand-specific information (unstranded)
- --tmo
- reports only those alignments within known transcriptome
- --dta
- reports alignments tailored for transcript assemblers
- --dta-cufflinks
- reports alignments tailored specifically for cufflinks
- --avoid-pseudogene
- tries to avoid aligning reads to pseudogenes (experimental option)
- --no-templatelen-adjustment
- disables template length adjustment for RNA-seq reads
- Scoring:
- --mp
<int>,<int>
- max and min penalties for mismatch; lower qual = lower penalty
<6,2>
- --sp
<int>,<int>
- max and min penalties for soft-clipping; lower qual = lower penalty
<2,1>
- --no-softclip
- no soft-clipping
- --np <int>
- penalty for non-A/C/G/Ts in read/ref (1)
- --rdg
<int>,<int>
- read gap open, extend penalties (5,3)
- --rfg
<int>,<int>
- reference gap open, extend penalties (5,3)
- --score-min
<func> min acceptable alignment score w/r/t read length
- (L,0.0,-0.2)
- Reporting:
- -k <int>
- It searches for at most <int> distinct, primary alignments for each
read. Primary alignments mean alignments whose alignment score is equal to
or higher than any other alignments. The search terminates when it cannot
find more distinct valid alignments, or when it finds <int>,
whichever happens first. The alignment score for a paired-end alignment
equals the sum of the alignment scores of the individual mates. Each
reported read or pair alignment beyond the first has the SAM
???secondary??? bit (which equals 256) set in its FLAGS field. For reads
that have more than <int> distinct, valid alignments, hisat2 does
not guarantee that the <int> alignments reported are the best
possible in terms of alignment score. Default: 5 (linear index) or 10
(graph index). Note: HISAT2 is not designed with large values for
-k in mind, and when aligning reads to long, repetitive genomes,
large -k could make alignment much slower.
- --max-seeds
<int>
- HISAT2, like other aligners, uses seed-and-extend approaches. HISAT2 tries
to extend seeds to full-length alignments. In HISAT2, --max-seeds
is used to control the maximum number of seeds that will be extended. For
DNA-read alignment (--no-spliced-alignment), HISAT2 extends up to
these many seeds and skips the rest of the seeds. For RNA-read alignment,
HISAT2 skips extending seeds and reports no alignments if the number of
seeds is larger than the number specified with the option, to be
compatible with previous versions of HISAT2. Large values for
--max-seeds may improve alignment sensitivity, but HISAT2 is not
designed with large values for --max-seeds in mind, and when
aligning reads to long, repetitive genomes, large --max-seeds could
make alignment much slower. The default value is the maximum of 5 and the
value that comes with -k times 2.
- -a/--all
- HISAT2 reports all alignments it can find. Using the option is equivalent
to using both --max-seeds and -k with the maximum value that
a 64-bit signed integer can represent (9,223,372,036,854,775,807).
- --repeat
- report alignments to repeat sequences directly
- Paired-end:
- -I/--minins
<int>
- minimum fragment length (0), only valid with
--no-spliced-alignment
- -X/--maxins
<int>
- maximum fragment length (500), only valid with
--no-spliced-alignment
--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
- Output:
- -t/--time
- print wall-clock time taken by search phases
- --un <path>
- write unpaired reads that didn't align to <path>
- --al <path>
- write unpaired reads that aligned at least once to <path>
- --un-conc
<path>
- write pairs that didn't align concordantly to <path>
- --al-conc
<path>
- write pairs that aligned concordantly at least once to <path>
- (Note: for --un, --al, --un-conc, or
--al-conc, add '-gz' to the option name, e.g. --un-gz
<path>, to gzip compress output, or add '-bz2' to bzip2 compress
output.) --summary-file <path> print alignment summary to
this file. --new-summary print alignment summary in a new style,
which is more machine-friendly. --quiet print nothing to stderr
except serious errors --met-file <path> send metrics to file
at <path> (off) --met-stderr send metrics to stderr (off)
--met <int> report internal counters & metrics every
<int> secs (1) --no-head suppress header lines, i.e. lines
starting with @ --no-sq suppress @SQ header lines --rg-id
<text> set read group id, reflected in @RG line and RG:Z: opt field
--rg <text> add <text> ("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.
- Performance:
-o/--offrate <int> override offrate of index; must
be >= index's offrate
-p/--threads <int> 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 'hisat2's can share
- Other:
- --qc-filter
- filter out reads that are bad according to QSEQ filter
- --seed
<int>
- seed for random number generator (0)
--non-deterministic seed rand. gen. arbitrarily instead
of using read attributes
- --remove-chrname
- remove 'chr' from reference names in alignment
- --add-chrname
- add 'chr' to reference names in alignment
- --version
- print version information and quit
- -h/--help
- print this usage message
64-bit Built on Debian 05 August 2021 Compiler: gcc version 10.2.1
20210110 (Debian 10.2.1-6) Options: -O3 -funroll-loops
-g3 -Wdate-time -D_FORTIFY_SOURCE=2
-std=c++11 Sizeof {int, long, long long, void*, size_t,
off_t}: {4, 8, 8, 8, 8, 8}