fasta3x.me

序列对齐 Compare a protein sequence to a protein sequence database or a DNA sequence to a DNA sequenc

.ip 
\fC\-m 5\fP provides a combination of \fC\-m 4\fP and
\fC\-m 0. \fC\-m 6 provides \fC\-m 5\fP plus HTML formatting.
.ip "-m 9"
provides coordinates and scores with the best score information.
A simple "\fC -m 9\fP extends the normal best score information:
.(l
.ft C
The best scores are:                                      opt bits E(14548)
XURTG4 glutathione transferase (EC 2.5.1.18) 4 -   ( 219) 1248 291.7 1.1e-79
.ft P
.)l
to include the additional information (on the same line, separated by
a <tab>):
.(l
.ft C
%_id  %_gid   sw  alen  an0  ax0  pn0  px0  an1  ax1 pn1 px1 gapq gapl  fs
0.771 0.771 1248  218    1  218    1  218    1  218    1  219   0   0   0
.ft P
.)l
\fC -m 9c\fP provides additional information: an encoded alignment string.  Thus:
.(l I
.ft C
       10        20        30        40        50          60         70  
GT8.7  NVRGLTHPIRMLLEYTDSSYDEKRYTMGDAPDFDRSQWLNEKFKL--GLDFPNLPYL-IDGSHKITQ
       :.::  . :: ::  .   .:::         : .:    ::.:   .: : ..:.. :::  :..:
XURTG  NARGRMECIRWLLAAAGVEFDEK---------FIQSPEDLEKLKKDGNLMFDQVPMVEIDG-MKLAQ
               20        30                 40        50        60        
.ft P
.)l
would be encoded:
.(l
.ft C
=23+9=13-2=10-1=3+1=5
.ft P
.)l
The alignment encoding is with repect to the alignment, not the
sequences.  The coordinate of the alignment is given earlier in the
"\fC -m 9c\fP" line.
.ip "-m 10"
\fC\-m 10\fP is a new, parseable format for use
with other programs.  See the file "readme.v20u4" for a more complete
description. 
.ip
As of version "fa34t23b2", it has become possible to combine independent
"\fC\-m\fP" options.  Thus, one can use "\fC\-m 1 -m 6 -m 9\fP".
.ip "-M low\-high"
Include library sequences (proteins only) with lengths between low and
high.
.ip "-n"
Force the query sequence to be treated as a DNA sequence.  This is
particularly useful for query sequences that contain a large number of
ambiguous residues, e.g. transcription factor binding sites.
.ip "-O"
Send copy of results to "filename."  Helpful for environments without
STDOUT (mostly for the Macintosh).
.ip "-o "
Turn off default optimization of all scores greater than OPTCUT. Sort
results by "initn" scores (reduces the accuracy of statistical
estimates).
.ip "-p"
Force query to be treated as protein sequence.
.ip "-Q,-q"
Quiet - does not prompt for any input.  Writes scores and alignments
to the terminal or standard output file.
.ip "-r"
Specify match/mismatch scores for DNA comparisons.  The default is
"+5/-4". "+3/-2" can perform better in some cases.
.ip "-R file"
Save a results summary line for every sequence in the sequence
library.  The summary line includes the sequence identifier,
superfamily number (if available) position 
in the library, and the similarity scores calculated.  This option can
be used to evaluate the sensitivity and selectivity of different
search strategies.[.wrp951,wrp981.]
.ip "-s file"
Specify the scoring matrix file.  \fCfasta3\fP uses the same scoring
matrices as Blast1.4/2.0.  Several scoring matrix files are included
in the standard distribution.  For protein sequences: \fCcodaa.mat\fP
- based on minimum mutation matrix; \fCidnaa.mat\fP - identity matrix;
\fCpam250.mat\fP - the PAM250 matrix developed by Dayhoff et
al.;[.day787.]  \fCpam120.mat\fP - a PAM120 matrix.  The default
scoring matrix is BLOSUM50 ("-s BL50"). Other matrices available from
within the program are: PAM250/"-s P250", PAM120/"-s P120", PAM40/"-s
P40", PAM20/"-s P20", MDM10 - MDM40/"-s M10 \- M40" (MDM are modern
PAM matrices from Jones et al.,[.tay925.]), BLOSUM50, 62, and 80/"-s
BL50", "-s BL62", "-s BL80".
.ip "-S"
Treat lower-case characters in the query or library sequences as
"low-complexity" ("seg"-ed) residues.  Traditionally, the "seg"
program [.woo935.] is used to remove low complexity regions in DNA
sequences by replacing the residues with an "X".  When the "-S" option
is used, the FASTA33 (and later) programs provide a potentially more
informative approach.  With "-S", lower case characters in the query
or database sequences are treated as "X"'s during the initial scan,
but are treated as normal residues during the final alignment display.
Since statistical significance is calculated from the similarity score
calculated during the library search, when the lower case residues are
"X"'s, low complexity regions will not produce statistically
significant matches.  However, if a significant alignment contains low
complexity regions, their alignmen is shown.  With "-S", lower case
characters may be included in the alignment to indicate low complexity
regions, and the final alignment score may be higher than the score
obtained during the search.
.ip
The \fCpseg\fP program can be used to produce databases (or query
sequences) with lower case residues indicating low complexity regions
using the command:
.(l I
\fCpseg database.fasta -z 1 -q  > database.lc_seg\fP
.)l
(\fCseg\fP can also be used with some post processing, see readme.v33tx.)
.ip
The \fC-S\fP option should always be used with \fCFASTX/Y\fCP and
\fCTFASTX/Y\fP because out of frame translations often generate
low-complexity protein sequences.  However, only lower case characters
in the protein sequence (or protein database) are masked; lower case
DNA sequences are translated into upper case protein sequences, and
not treated as low complexity by the translated alignment programs.
.ip "-t #"
Translation table - tfasta3, fastx3, tfastx3, fasty3, and
tfasty3 now support the BLAST tranlation tables.  See
\fChttp://www.ncbi.nlm.nih.gov/Taxonomy/Utils/wprintgc.cgi\fP.
.ip
In addition, "\-t t" or "\-t t#" turns on the addition of an implicit termination
codon to a protein:translated DNA match.  That is, each protein
sequence implicitly ends with "*", which matches the termination codes
for the appropriate genetic code.  "\-t t#" sets implicit termination
and a different genetic code.
.ip "-U"
Treat the query sequence an RNA sequence.  In addition to selecting a
DNA/RNA alphabet, this option causes changes to the scoring matrix so
that 'G:A' , 'T:C' or 'U:C' are scored as 'G:G'.
.ip "-V str"
It is now possible to specify some annotation characters that can be
included (and will be ignored), in the query sequence file.  Thus, One
might have a file with: \fC"ACVS*ITRLFT?"\fP, where "*" and "?"  are
used to indicate phosphorylation.  By giving the option \fC\-V '*?'\fP,
those characters in the query will be moved to an "annotation string",
and alignments that include the annotated residues will be highlighted
with the appropriate character above the sequence (on the number line).
.ip "-w #"
Line length (width) = number (<200)
.ip "-W #"
 context length (default is 1/2 of line width -w) for alignment,
like fasta and ssearch, that provide additional sequence context.
.ip "-x #match,#mismatch"
Specify the penalty for a match to an 'X', and mismatch to 'X',
independently of the PAM matrix.  Particularly useful for
\fCfastx3/fasty3\fP, where termination codons are encoded as 'X'.
.ip "-X \"off1 off2\""
Specifies offsets for the beginning of the query and library sequence.
For example, if you are comparing upstream regions for two genes, and
the first sequence contains 500 nt of upstream sequence while the
second contains 300 nt of upstream sequence, you might try:
.(l I
\fCfasta -X "-500 -300" seq1.nt seq2.nt\fP
.)l
If the -X option is not used, FASTA assumes numbering starts with 1.
(You should double check to be certain the negative numbering works
properly.)
.ip "-y"
Set the width of the band used for calculating "optimized" scores.
For proteins and ktup=2, the width is 16.  For proteins with ktup=1,
the width is 32 by default.  For DNA the width is 16.
.ip "-z -1,0,1,2,3,4,5"
\fC\-z -1\fP turns off statistical calculations. \fCz 0\fP estimates
the significance of the match from the mean and standard deviation of
the library scores, without correcting for library sequence length.
\fC\-z 1\fP (the default) uses a weighted regression of average score
vs library sequence length; \fC\-z 2\fP uses maximum likelihood
estimates of
.if t \(*l
.if n Lambda
and \fIK\fP; \fC\-z 3\fP uses Altschul-Gish
parameters;[.alt960.] \fC\-z 4 \- 5\fP uses two variations on the
\fC\-z 1\fP strategy. \fC\-z 1\fP and \fC\-z 2\fP are the best methods,
in general.
.ip "-z 11,12,14,15" 
estimate the statistical parameters from shuffled copies of each
library sequence.  This doubles the time required for a search, but
allows accurate statistics to be estimated for libraries comprised of
a single protein family.
.ip "-Z db_size"
set the apparent size of the database to be used when calculating
expectation E() values.  If you searched a database with 1,000
sequences, but would like to have the E()-values calculated in the
context of a 100,000 sequence database, use '-Z 100000'.
.ip "-1"
sort output by init1 score (for compatibility with FASTP - do not
use).
.ip "-3"
translate only three forward frames
.sp
.lp
For example:
.(l
\fCfasta -w 80 -a seq1.aa seq.aa\fP
.)l
would compare the sequence in seq1.aa to that in seq2.aa and display the
results with 80 residues on an output line, showing all of the residues
in both sequences.  Be sure to enter the options before entering the file
names, or just enter the options on the command line, and the program will
prompt for the file names.
.sp
.pp
(November, 1997) In addition, it is now possible to provide the fasta
programs with the query sequence (fasta, fasty, ssearch, tfastx), or
two sequences (prss, lalign, plalign) from the unix "stdin" stream.  This
makes it much easier to set up FASTA or PRSS WWW pages.  To specify
that stdin be used, rather than a file, the file name should be
specified as '-' or '@' (the latter file name makes it possible to
specify a subset of the sequence).
Thus:
.(l
cat query.aa | fasta -q @:25-75 s
.)l
would take residues 25-75 from query.aa and search the 's' library
(see the discussion of FASTLIBS).
.sh 2 "Environment variables"
.pp
Because the current version of the program allows the user to set
virtually every option on the command line (except the \fIktup\fP,
which must be set as the third command line argument), only the
\fCFASTLIBS\fP environment variable is routinely used.
.ip "FASTLIBS"
specifies the location of the file which contains the list of library
descriptions, locations, and library types (see section on finding
library files).
.sh 1 "Frequently Asked Questions (FAQs)"
.np
\fIWhich program should I use?\fP See Table I.
.np
\fIHow do I search with both DNA strands with\fP \fCfasta3\fP \fIand\fP
\fCfastx3\fP? With version 32 of the FASTA program package, all
searches that use DNA queries (e.g. \fCfasta3\fP, \fCfastx3/y3\fP)
examine both strands. To revert to earlier FASTA behavior - only
looking at the forward or reverse strand - use \fC\-3\fP to search only
the forward strand and \fC\-i -3\fP to search only the reverse strand.
.np
\fIWhen I search Genbank - the program reports:\fP \fC0 residues in 0
sequences\fP.  This typically happens because the program does not
know that you are searching a Genbank flatfile database and is looking
for a FASTA format database.  Be certain to specify the library type
("1" for Genbank flatfile) with the database name.
.np
What is the difference between \fCfastx3\fP and \fCfasty3\fP (or
\fCtfastx3\fP and \fCtfasty3\fP).  \fC[t]fastx3\fP uses a simpler
codon based model for alignments that does not allow frameshifts in
some codon positions (see ref. [.wrp971.]).  \fCtfastx3\fP is about
30% faster, but \fCtfasty3\fP can produce higher quality alignments in
some cases.
.np
\fIWhen I run\fP \fCfasta3 -q\fP, I don't see any (or very little)
output, but I get lots of scores when I run interactively. With the
\fC\-Q\fP option, the number of high scores displayed is limited by the
\fC\-E #\fP cutoff, which is 10.0 for protein comparisons, 2.0 for DNA
comparisons, and 5.0 for translated DNA:protein comparisons.  In
interactive mode (without \fC\-Q\fP), by default you see 20 high
scores, regardless of \fCE()\fP value.
.np
\fIWhat is ktup\fP \- All of the programs with \fCfast\fP in their
name use a computer science method called a lookup table to speed the
search.  For proteins with \fIktup\fP=2, this means that the program
does not look at any sequence alignment that does not involve matching
two identical residues in both sequences.  Likewise with DNA and
\fIktup\fP = 6, the initial alignment of the sequences looks for 6
identical adjacent nucleotides in both sequences.  Because it is less
likely that two identical amino-acids will line up by chance in two
unrelated proteins, this speeds up the comparison.  But very distantly
related sequences may never have two identical residues in a row but
will have single aligned identities.  In this case, \fIktup\fP = 1 may
find alignments that \fIktup\fP=2 misses.
.np
\fISometimes, in the list of best scores, the same sequence is shown
twice with exactly the same score.  Sometimes, the sequence is there
twice, but the scores are slightly different.\fP When any of the
\fCfasta3\fP programs searches a long sequence, it breaks the sequence
up into \fIoverlapping\fP pieces.  The length of the piece depends on
the length of the query and the particular program being used (it can
also be controlled with the -N #### option).  Since the pieces overlap
by the length of the query sequence (or 3*query_length for fastx/y3
and tfasta/x/y3), if the highest scoring alignment is at the end of
one piece, it will be scored again at the beginning of the next piece.
If the alignment is not be completely included in the overlap region,
one of the pieces will give a higher score than the other.  These
duplications can be detected by looking at the coordinates of the
alignment.  If either the beginning or end coordinate is identical in
two alignments, the alignments are at least partially duplicates.
.lp
As always, please inform me of bugs as soon as possible.
.sp
.nf
William R. Pearson
Department of Biochemistry
Jordan Hall Box 800733
U. of Virginia
Charlottesville, VA

[e-mail]

.sh 1 "References"
.[]