Ubuntu Manpage: nhmmer - search DNA queries against a DNA sequence database

NAME

       nhmmer - search DNA queries against a DNA sequence database

SYNOPSIS

       nhmmer [options] queryfile seqdb

DESCRIPTION

nhmmer is used to search one or more nucleotide queries against a nucleotide sequence
database. For each query in queryfile, use that query to search the target database of
sequences in seqdb, and output a ranked list of the hits with the most significant matches
to the query. A query may be either a profile model built using hmmbuild, a sequence
alignment, or a single sequence. Sequence based queries can be in a number of formats (see
--qformat), and can typically be autodetected. Note that only Stockholm format supports
queries made up of more than one sequence alignment.

Either the query queryfile or the target seqdb may be '-' (a dash character), in which
case the query file or target database input will be read from a <stdin> pipe instead of
from a file. Only one input source can come through <stdin>, not both. If the queryfile
contains more than one query, then seqdb cannot come from stdin, because we can't rewind
the streaming target database to search it with another profile.

If the query is sequence-based (unaligned or aligned), a new file containing the HMM(s)
built from the input(s) in queryfile may optionally be produced, with the filename set
using the --hmmout flag.

The output format is designed to be human-readable, but is often so voluminous that
reading it is impractical, and parsing it is a pain. The --tblout option saves output in a
simple tabular format that is concise and easier to parse. The -o option allows
redirecting the main output, including throwing it away in /dev/null.

OPTIONS

       -h     Help; print a brief reminder of command line usage and all available options.

OPTIONS FOR CONTROLLING OUTPUT

       -o <f> Direct the main human-readable output to a file <f> instead of the default stdout.

       -A <f> Save a multiple alignment of all  significant  hits  (those  satisfying  "inclusion
              thresholds") to the file <f>.

       --tblout <f>
              Save  a  simple  tabular  (space-delimited) file summarizing the per-target output,
              with one data line per homologous target sequence found.

       --dfamtblout <f>
              Save a tabular (space-delimited) file summarizing the per-hit  output,  similar  to
              --tblout but more succinct.

       --aliscoresout <f>
              Save  to  file  a  list  of  per-position scores for each hit.  This is useful, for
              example, in identifying  regions  of  high  score  density  for  use  in  resolving
              overlapping hits from different models.

       --hmmout <f>
              If queryfile is sequence-based, write the internally-computed HMM(s) to file <f>.

       --acc  Use  accessions  instead  of names in the main output, where available for profiles
              and/or sequences.

       --noali
              Omit the alignment section from the main output. This can greatly reduce the output
              volume.

       --notextw
              Unlimit  the  length of each line in the main output. The default is a limit of 120
              characters per line, which helps in displaying the output cleanly on terminals  and
              in editors, but can truncate target profile description lines.

       --textw <n>
              Set  the main output's line length limit to <n> characters per line. The default is
              120.

OPTIONS CONTROLLING SINGLE SEQUENCE SCORING

By default, if a query is a single sequence from a file in fasta format, nhmmer uses a
search model constructed from that sequence and a standard 20x20 substitution matrix for
residue probabilities, along with two additional parameters for position-independent gap
open and gap extend probabilities. These options allow the default single-sequence scoring
parameters to be changed, and for single-sequence scoring options to be applied to a
single sequence coming from an aligned format.

--singlemx
If a single sequence query comes from a multiple sequence alignment file, such as
in Stockholm format, the search model is by default constructed as is typically
done for multiple sequence alignments. This option forces nhmmer to use the single-
sequence method with substitution score matrix.

--mxfile<mxfile
Obtain residue alignment probabilities from the substitution matrix in file mxfile.
The default score matrix is DNA1 (this matrix is internal to HMMER and does not
have to be available as a file). The format of a substitution matrix mxfile is the
standard format accepted by BLAST, FASTA, and other sequence analysis software.
See ftp.ncbi.nlm.nih.gov/blast/matrices/ for example files. (The only exception: we
require matrices to be square, so for DNA, use files like NCBI's NUC.4.4, not
NUC.4.2.)

--popen <x>
Set the gap open probability for a single sequence query model to <x>. The default
is 0.02. <x> must be >= 0 and < 0.5.

--pextend <x>
Set the gap extend probability for a single sequence query model to <x>. The
default is 0.4. <x> must be >= 0 and < 1.0.

OPTIONS CONTROLLING REPORTING THRESHOLDS

       Reporting thresholds control which hits are reported in output  files  (the  main  output,
       --tblout, and --dfamtblout).  Hits are ranked by statistical significance (E-value).

       -E <x> Report  target  sequences  with an E-value of <= <x>.  The default is 10.0, meaning
              that on average, about 10 false positives will be reported per query,  so  you  can
              see the top of the noise and decide for yourself if it's really noise.

       -T <x> Instead  of  thresholding output on E-value, instead report target sequences with a
              bit score of >= <x>.

OPTIONS FOR INCLUSION THRESHOLDS

       Inclusion thresholds are stricter than reporting thresholds.  Inclusion thresholds control
       which hits are considered to be reliable enough to be included in an output alignment or a
       subsequent search round, or marked as significant ("!") as opposed to  questionable  ("?")
       in hit output.

       --incE <x>
              Use  an E-value of <= <x> as the inclusion threshold.  The default is 0.01, meaning
              that on average, about 1 false positive would be expected  in  every  100  searches
              with different query sequences.

       --incT <x>
              Instead  of  using E-values for setting the inclusion threshold, use a bit score of
              >= <x> as the inclusion threshold.  By default this option is unset.

OPTIONS FOR MODEL-SPECIFIC SCORE THRESHOLDING

       Curated profile databases may define specific  bit  score  thresholds  for  each  profile,
       superseding any thresholding based on statistical significance alone.

       To  use  these  options,  the  profile  must  contain  the appropriate (GA, TC, and/or NC)
       optional score threshold annotation; this is picked up by hmmbuild from  Stockholm  format
       alignment  files.  For  a  nucleotide model, each thresholding option has a single per-hit
       threshold <x> This acts as if -T <x> --incT <x> has been applied specifically  using  each
       model's curated thresholds.

       --cut_ga
              Use  the  GA  (gathering) bit score threshold in the model to set per-hit reporting
              and inclusion thresholds. GA thresholds are generally considered to be the reliable
              curated  thresholds  defining  family  membership;  for  example,  in  Dfam,  these
              thresholds are applied when annotating a genome with a model of a family  known  to
              be  found  in  that  organism.  They may allow for minimal expected false discovery
              rate.

       --cut_nc
              Use the NC (noise cutoff) bit score threshold in the model to set per-hit reporting
              and  inclusion thresholds. NC thresholds are less stringent than GA; in the context
              of Pfam, they are generally used to store the score of  the  highest-scoring  known
              false positive.

       --cut_tc
              Use  the  TC  (trusted  cutoff)  bit  score  threshold  in the model to set per-hit
              reporting and inclusion thresholds. TC thresholds are more stringent than  GA,  and
              are  generally considered to be the score of the lowest-scoring known true positive
              that is above all known false positives; for example, in Dfam, these thresholds are
              applied  when annotating a genome with a model of a family not known to be found in
              that organism.

OPTIONS CONTROLLING THE ACCELERATION PIPELINE

HMMER3 searches are accelerated in a three-step filter pipeline: the scanning-SSV filter,
the Viterbi filter, and the Forward filter. The first filter is the fastest and most
approximate; the last is the full Forward scoring algorithm. There is also a bias filter
step between SSV and Viterbi. Targets that pass all the steps in the acceleration pipeline
are then subjected to postprocessing -- domain identification and scoring using the
Forward/Backward algorithm.

Changing filter thresholds only removes or includes targets from consideration; changing
filter thresholds does not alter bit scores, E-values, or alignments, all of which are
determined solely in postprocessing.

--max Turn off (nearly) all filters, including the bias filter, and run full
Forward/Backward postprocessing on most of the target sequence. In contrast to
phmmer and hmmsearch, where this flag really does turn off the filters entirely,
the --max flag in nhmmer sets the scanning-SSV filter threshold to 0.4, not 1.0.
Use of this flag increases sensitivity somewhat, at a large cost in speed.

--F1 <x>
Set the P-value threshold for the SSV filter step. The default is 0.02, meaning
that roughly 2% of the highest scoring nonhomologous targets are expected to pass
the filter.

--F2 <x>
Set the P-value threshold for the Viterbi filter step. The default is 0.001.

--F3 <x>
Set the P-value threshold for the Forward filter step. The default is 1e-5.

--nobias
Turn off the bias filter. This increases sensitivity somewhat, but can come at a
high cost in speed, especially if the query has biased residue composition (such as
a repetitive sequence region, or if it is a membrane protein with large regions of
hydrophobicity). Without the bias filter, too many sequences may pass the filter
with biased queries, leading to slower than expected performance as the
computationally intensive Forward/Backward algorithms shoulder an abnormally heavy
load.

OPTIONS FOR SPECIFYING THE ALPHABET

       --dna  Assert that sequences in msafile are DNA, bypassing alphabet autodetection.

       --rna  Assert that sequences in msafile are RNA, bypassing alphabet autodetection.

OPTIONS CONTROLLING SEED SEARCH HEURISTIC

When searching with nhmmer, one may optionally precompute a binary version of the target
database, using makehmmerdb, then search against that database. Using default settings,
this yields a roughly 10-fold acceleration with small loss of sensitivity on benchmarks.
This is achieved using a heuristic method that searches for seeds (ungapped alignments)
around which full processing is done. This is essentially a replacement to the SSV stage.
(This method has been extensively tested, but should still be treated as somewhat
experimental.) The following options only impact nhmmer if the value of --tformat is
hmmerdb.

Changing parameters for this seed-finding step will impact both speed and sensitivity -
typically faster search leads to lower sensitivity.

--seed_max_depth <n>
The seed step requires that a seed reach a specified bit score in length no longer
than <n>. By default, this value is 15. Longer seeds allow a greater chance of
meeting the bit score threshold, leading to diminished filtering (greater
sensitivity, slower run time).

--seed_sc_thresh <x>
The seed must reach score <x> (in bits). The default is 15.0 bits. A higher
threshold increases filtering stringency, leading to faster run times and lower
sensitivity.

--seed_sc_density <x>
Either all prefixes or all suffixes of a seed must have bit density (bits per
aligned position) of at least <x>. The default is 0.8 bits/position. An increase
in the density requirement leads to increased filtering stringency, thus faster run
times and lower sensitivity.

--seed_drop_max_len <n>
A seed may not have a run of length <n> in which the score drops by --seed_drop_lim
or more. Basically, this prunes seeds that go through long slightly-negative seed
extensions. The default is 4. Increasing the limit causes (slightly) diminished
filtering efficiency, thus slower run times and higher sensitivity. (minor tuning
option)

--seed_drop_lim <x>
In a seed, there may be no run of length --seed_drop_max_len in which the score
drops by --seed_drop_lim. The default is 0.3 bits. Larger numbers mean less
filtering. (minor tuning option)

--seed_req_pos <n>
A seed must contain a run of at least <n> positive-scoring matches. The default is
5. Larger values mean increased filtering. (minor tuning option)

--seed_ssv_length <n>
After finding a short seed, an ungapped alignment is extended in both directions in
an attempt to meet the --F1 score threshold. The window through which this ungapped
alignment extends is length <n>. The default is 70. Decreasing this value
slightly reduces run time, at a small risk of reduced sensitivity. (minor tuning
option)

OTHER OPTIONS

--qformat <s>
Assert that input queryfile is a sequence file (unaligned or aligned), in format
<s>, bypassing format autodetection. Common choices for <s> include: fasta, embl,
genbank. Alignment formats also work, and will serve as the basis for automatic
creation of a profile HMM used for searching; common choices include: stockholm,
a2m, afa, psiblast, clustal, phylip. For more information, and for codes for some
less common formats, see main documentation.

--qsingle_seqs
Force queryfile to be read as individual sequences, even if it is in an msa format.
For example, if the input is in aligned stockholm format, the --qsingle_seqs
flag will cause each sequence in that alignment to be used as a separate query
sequence.

--tformat <s>
Assert that target sequence database seqdb is in format <s>, bypassing format
autodetection. Common choices for <s> include: fasta, embl, genbank, ncbi,
fmindex. Alignment formats also work; common choices include: stockholm, a2m, afa,
psiblast, clustal, phylip. For more information, and for codes for some less
common formats, see main documentation. The string <s> is case-insensitive (fasta
or FASTA both work). The format ncbi indicates that the database file is a binary
file produced using makeblastdb. The format fmindex indicates that the database
file is a binary file produced using makehmmerdb.

--nonull2
Turn off the null2 score corrections for biased composition.

-Z <x> For the purposes of per-hit E-value calculations, Assert that the total size of the
target database is <x> million nucleotides, rather than the actual number of
targets seen.

--seed <n>
Set the random number seed to <n>. Some steps in postprocessing require Monte
Carlo simulation. The default is to use a fixed seed (42), so that results are
exactly reproducible. Any other positive integer will give different (but also
reproducible) results. A choice of 0 uses a randomly chosen seed.

--w_beta <x>
Window length tail mass. The upper bound, W, on the length at which nhmmer expects
to find an instance of the model is set such that the fraction of all sequences
generated by the model with length >= W is less than <x>. The default is 1e-7.
This flag may be used to override the value of W established for the model by
hmmbuild, or when the query is sequence-based.

--w_length <n>
Override the model instance length upper bound, W, which is otherwise controlled by
--w_beta. It should be larger than the model length. The value of W is used deep
in the acceleration pipeline, and modest changes are not expected to impact results
(though larger values of W do lead to longer run time). This flag may be used to
override the value of W established for the model by hmmbuild, or when the query is
sequence-based.

--watson
Only search the top strand. By default both the query sequence and its reverse-
complement are searched.

--crick
Only search the bottom (reverse-complement) strand. By default both the query
sequence and its reverse-complement are searched.

--cpu <n>
Set the number of parallel worker threads to <n>. On multicore machines, the
default is 2. You can also control this number by setting an environment variable,
HMMER_NCPU. There is also a master thread, so the actual number of threads that
HMMER spawns is <n>+1.

This option is not available if HMMER was compiled with POSIX threads support
turned off.

--stall
For debugging the MPI master/worker version: pause after start, to enable the
developer to attach debuggers to the running master and worker(s) processes. Send
SIGCONT signal to release the pause. (Under gdb: (gdb) signal SIGCONT) (Only
available if optional MPI support was enabled at compile-time.)

--mpi Run under MPI control with master/worker parallelization (using mpirun, for
example, or equivalent). Only available if optional MPI support was enabled at
compile-time.

COPYRIGHT

       Copyright (C) 2020 Howard Hughes Medical Institute.
       Freely distributed under the BSD open source license.

       For  additional  information  on copyright and licensing, see the file called COPYRIGHT in
       your HMMER source distribution, or see the HMMER web page (http://hmmer.org/).

AUTHOR

       http://eddylab.org