FASTX+

Table of Contents

FUNCTION

DESCRIPTION

EXAMPLE

FUNCTION

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FastX+ does a Pearson and Lipman search for similarity between a nucleotide query sequence and a group of protein sequences, taking frameshifts into account. FastX+ translates both strands of the nucleic sequence before performing the comparison. It is designed to answer the question, "What implied protein sequences in my nucleic acid sequence are similar to sequences in a protein database?"

DESCRIPTION

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Advantages of Plus “+” Programs:

P Plus programs are enhanced to be able to read sequences in a variety of native formats such as GCG RSF, GCG SSF, GCG MSF, GenBank, EMBL, FastA, SwissProt, PIR, and BSML without conversion.

P Plus programs remove sequence length restriction of 350,000bp.

If you do not need these features and wish to have more interactivity, you might wish to seek out and run the original program version.

FastX+ uses the method of Pearson and Lipman (Proc. Natl. Acad. Sci. USA 85; 2444-2448 (1988)) to search for similarities between a nucleic acid sequence (the query) and any group of protein sequences. Like FrameSearch, it is especially useful when the nucleotide query contains sequencing errors that cause frameshifts that would flaw the translated sequence so that it wouldn't find matches using a protein-protein search method such as FastA+. FastX+ is faster than FrameSearch, but not as sensitive.

In the first step of this search, the comparison can be viewed as a set of dot plots, with the query as the vertical sequence and the group of sequences to which the query is being compared as the different horizontal sequences. This first step finds the registers of comparison (diagonals) having the largest number of short perfect matches (words) for each comparison. In the second step, these "best" regions are rescored using a scoring matrix that allows conservative replacements, ambiguity symbols, and runs of identities shorter than the size of a word. In the third step, the program checks to see if some of these initial highest-scoring diagonals can be joined together. Finally, the search set sequences with the highest scores are aligned to the query sequence for display.

What is a Word?

A word is any short sequence (n-mer or k-tuple) where you have set n to some small integer less than or equal to six. The word GGATGG is one of the 4,096 possible words of length six that can be created from an alphabet consisting of the four letters G, A, T, and C. The word QL is one of the 400 possible words of length two that you can make with the 20 letters of the amino acid alphabet.

EXAMPLE

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Here is a session using FastX+ to identify sequences in the Uniprot protein sequence database that are similar to a fragment of an actinomyces species coding region:

17:17~195> fastx+ -histogram=true

FastX+ does a Pearson and Lipman search for similarity between a nucleotide query sequence and a group of protein sequences, taking frameshifts into account. FastX+ translates both strands of the nucleic sequence before performing the comparison. It is designed to query a nucleotide sequence against a protein database by doing a six frame translation. It answers the question, "What implied protein sequences in my nucleic acid sequence are similar to sequences in a protein database?"

fastx+ with what query sequence(s) ? gb_ba:a16stm210

Begin (* 1 *) ?

End (-1 for entire sequence) (* -1 *) ? 200

Enter value for search set (*Default DB*) ? uniprot:*

What should I call the output file (* <sequence_name>.<program_name> *) ?

Creating A16STM210.fastx+ as the output file.

OUTPUT

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The output from FastX+ is a list file, and is suitable for input to any GCG program that allows indirect file specifications. (For information about indirect file specification, see Section 2, Using Sequence Files and Databases of the User's Guide.)

Here is some of the output file:

# $GCGROOT/bin/fastx34_native -O /var/tmp/bslskAAATNa4Bh.tmp -E 2.0 -b 10 -T 1 /var/tmp/bslskBAAVNa4Bh.fa "uniprot:* 17"

 FASTX compares a DNA sequence to a protein sequence data bank

 version 3.4t21 May 14, 2003

Please cite:

 Pearson et al, Genomics (1997) 46:24-36

Query library /var/tmp/bslskBAAVNa4Bh.fa vs uniprot:* library

searching uniprot:* 17 library

  1>>>A16STM210 X92704 Actinomyces species 16S ribosomal RNA (isolate TM210). 10/1996 - 200 aa

 vs  uniprot:* library

       opt      E()

< 20  1420     0:=

  22     5     0:=          one = represents 2446 library sequences

  24    26     1:*

  26    60    32:*

  28   275   341:*

  30  1394  2073:*

  32  4838  8015:== *

  34 15010 21735:======= *

  36 39665 44639:================= *

  38 73671 73771:==============================*

  40 118004 102904:==========================================*======

  42 135840 125788:===================================================*====

  44 146701 138756:========================================================*===

  46 140657 141326:=========================================================*

  48 129284 135304:=====================================================  *

  50 116199 123465:================================================  *

  52 98570 108546:=========================================   *

  54 81451 92718:==================================   *

  56 68806 77448:=============================  *

  58 61017 63583:=========================*

  60 55350 51506:=====================*=

  62 47218 41293:================*===

  64 37431 32840:=============*==

  66 30578 25956:==========*==

  68 23604 20416:========*=

  70 18513 15999:======*=

  72 14357 12502:=====*

  74 11070  9747:===*=

  76  8541  7587:===*

  78  6841  5897:==*

  80  4602  4579:=*

  82  3610  3503:=*

  84  2647  2775:=*

  86  1838  2147:*

  88  1405  1661:*          inset = represents 17 library sequences

  90  1179  1285:*

  92   833   994:*         :=======================================*

  94   558   769:*         :=================================      *

  96   456   595:*         :===========================       *

  98   320   461:*         :===================        *

 100   199   356:*         :============        *

 102   132   276:*         :========        *

 104    99   213:*         :======      *

 106    81   165:*         :=====    *

 108    63   128:*         :====   *

 110    38    99:*         :===  *

 112    37    76:*         :=== *

 114    15    59:*         :=  *

 116    13    46:*         := *

 118     8    35:*         := *

>120    23    27:*         :=*

475286682 residues in 1504552 sequences

 statistics sampled from 60000 to 1504532 sequences

  Expectation_n fit: rho(ln(x))= 4.9258+/-0.00017; mu= 5.7905+/- 0.009

 mean_var=48.9373+/- 9.703, 0's: 60 Z-trim: 60  B-trim: 2222 in 1/65

 Lambda= 0.183339

 Kolmogorov-Smirnov  statistic: 0.0193 (N=29) at  58

FASTX (3.46 May 2003) function [optimized, BL50 matrix (o=15:-5:-1)] ktup: 2

 join: 36, opt: 30, open/ext: -14/-2 shift: -20, width:  16

 Scan time: 5033.050

The best scores are:                                       opt bits E(1504552)

Q09649 Q09649 caenorhabditis elegans. hypothet ( 702) [r]   99  34.0      12

Q7U714 Q7u714 synechococcus sp. (strain wh8102 ( 144) [f]   88  30.9      21

Q8LE84 Q8le84 arabidopsis thaliana (mouse-ear  ( 144) [r]   86  30.4      31

Q9PW96 Q9pw96 balearica pavonina gibbericeps.  ( 209) [f]   87  30.7      36

Q9PW92 Q9pw92 pachyptila turtur (fairy prion). ( 215) [f]   87  30.7      37

Q9PW91 Q9pw91 pachyptila desolata. prion prote ( 215) [f]   87  30.7      37

Q47572 Q47572 escherichia coli. plasmid r100 p ( 189) [r]   86  30.4      39

Q9I9F3 Q9i9f3 columba rupestris. prion protein ( 266) [f]   87  30.7      45

Q49087 Q49087 mycoplasma capricolum. adenylosu ( 120) [f]   83  29.6      45

AAR19695 Aar19695 ateles fusciceps (brown-head ( 225) [r]   85  30.2      55

200 residues in 1 query   sequences

475286682 residues in 1504552 library sequences

 Scomplib [34t21]

 start: Wed Dec  8 17:17:54 2004 done: Wed Dec  8 19:01:38 2004

 Total Scan time: 5033.050 Total Display time:  2.150

        Function used was FASTX [version 3.4t21 May 14, 2003]

What is the Output?

The first part of the output file contains a histogram showing the distribution of the z-scores between the query and search set sequences. The histogram is composed of bins of size 2 that are labeled according to the higher score for that bin (the leftmost column of the histogram). For example, the bin labeled 24 stores the number of sequence pairs that had scores of 23 or 24.

The next two columns of the histogram list the number of z-scores that fell within each bin. The second column lists the number of z-scores observed in the search and the third column lists the number of z-scores that were expected.

The body of the histogram displays a graphical representation of the score distributions. Equal signs (=) indicate the number of scores of that magnitude that were observed during the search, while asterisks (*) plot the number of scores of that magnitude that were expected.

At the bottom of the histogram is a list of some of the parameters pertaining to the search.

Below the histogram, FastX+ displays a listing of the best scores. Strand:- after the sequence name in this list indicates that the match was found between search set sequence and the reverse complement of the query sequence.

Following the list of best scores, FastX+ displays the alignments of the regions of best overlap between the query and search sequences. /rev following the query sequence name indicates that the search sequence is aligned with the reverse complement of the query sequence.

This program displays only the region of overlap between the two aligned sequences (plus some residues on either side of the region to provide context for the alignment) unless you use -showall. The display of identities and conservative replacements between the aligned sequences depends on the value of -markx. By default (-markx=3), the pipe character (|) is used to denote identities and the colon (:) to denote conservative replacements.

INPUT FILES

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FastX+ accepts a single nucleic acid sequence as the query sequence. The search set is either a single protein sequence or multiple protein sequences. You can specify multiple sequences in a number of ways: by using a list file, for example @project.list; by using an MSF or RSF file, for example project.msf{*}; or by using a sequence specification with an asterisk (*) wildcard, for example Genbank:*. If FastX+ rejects your nucleotide sequence, turn to Appendix VI to see how to change or set the type of a sequence.

RELATED PROGRAMS

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FastX does a Pearson and Lipman search for similarity between a nucleotide query sequence and a group of protein sequences, taking frameshifts into account. FastX translates both strands of the nucleic sequence before performing the comparison. It is designed to answer the question, "What implied protein sequences in my nucleic acid sequence are similar to sequences in a protein database?"

FastA+ does a Pearson and Lipman search for similarity between a query sequence and a group of sequences of the same type (nucleic acid or protein). For nucleotide searches, FastA+ may be more sensitive than BLAST+.

BLAST+ searches one or more nucleic acid or protein databases for sequences similar to one or more query sequences of any type. BLAST+ can produce gapped alignments for the matches it finds. NetBLAST searches for sequences similar to a query sequence. The query and the database searched can be either peptide or nucleic acid in any combination. NetBLAST+ can search only databases maintained at the National Center for Biotechnology Information (NCBI) in Bethesda, Maryland, USA.

SSearch+ does a rigorous Smith-Waterman search for similarity between a query sequence and a group of sequences of the same type (nucleic acid or protein). This may be the most sensitive method available for similarity searches. Compared to BLAST+ and FastA+, it can be very slow.

TFastA+ does a Pearson and Lipman search for similarity between a protein query sequence and any group of nucleotide sequences. TFastA+ translates the nucleotide sequences in all six reading frames before performing the comparison. It is designed to answer the question, "What implied protein sequences in a nucleotide sequence database are similar to my protein sequence?"

TFastX+ does a Pearson and Lipman search for similarity between a protein query sequence and any group of nucleotide sequences, taking frameshifts into account. It is designed to be a replacement for TFastA+, and like TFastA+, it is designed to answer the question, "What implied protein sequences in a nucleotide sequence database are similar to my protein sequence?"

FrameSearch searches a group of protein sequences for similarity to one or more nucleotide query sequences, or searches a group of nucleotide sequences for similarity to one or more protein query sequences. For each sequence comparison, the program finds an optimal alignment between the protein sequence and all possible codons on each strand of the nucleotide sequence. Optimal alignments may include reading frame shifts.

WordSearch identifies sequences in the database that share large numbers of common words in the same register of comparison with your query sequence. The output of WordSearch can be displayed with Segments.

ProfileSearch and MotifSearch use a profile (derived from a set of aligned sequences) instead of a query sequence to search a collection of sequences. FindPatterns+ uses a pattern described by a regular expression to search a collection of sequences.

StringSearch, LookUp, and Names identify sequences by searching the annotation (non-sequence) portions of seqence files or sequence databases.

FastA does a Pearson and Lipman search for similarity between a query sequence and a group of sequences of the same type (nucleic acid or protein). For nucleotide searches, FastA may be more sensitive than BLAST.

BLAST searches one or more nucleic acid or protein databases for sequences similar to one or more query sequences of any type. BLAST can produce gapped alignments for the matches it finds. NetBLAST searches for sequences similar to a query sequence. The query and the database searched can be either peptide or nucleic acid in any combination. NetBLAST can search only databases maintained at the National Center for Biotechnology Information (NCBI) in Bethesda, Maryland, USA.

SSearch does a rigorous Smith-Waterman search for similarity between a query sequence and a group of sequences of the same type (nucleic acid or protein). This may be the most sensitive method available for similarity searches. Compared to BLAST and FastA, it can be very slow.

TFastA does a Pearson and Lipman search for similarity between a protein query sequence and any group of nucleotide sequences. TFastA translates the nucleotide sequences in all six reading frames before performing the comparison. It is designed to answer the question, "What implied protein sequences in a nucleotide sequence database are similar to my protein sequence?"

TFastX does a Pearson and Lipman search for similarity between a protein query sequence and any group of nucleotide sequences, taking frameshifts into account. It is designed to be a replacement for TFastA, and like TFastA, it is designed to answer the question, "What implied protein sequences in a nucleotide sequence database are similar to my protein sequence?"

RESTRICTIONS

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The query sequence cannot be longer than 20,000 symbols. You cannot select a list size of more than 1,000 best scores nor view more than 1,000 alignments. The word size must be either 1 or 2.

For the estimates of statistical significance to be valid, the search set must contain a large sample of unrelated sequences. The statistical estimates will not be calculated at all if there are fewer than 10 sequences in the search set (20 sequences if only one strand is searched).

With -nooptall, the estimates of statistical significance will not be accurate.

Fast suite of programs work with the flat file databases only. Users cannot specify Blast databases as a database specification for FastX+.

For Tru64 (OSF) FastX+ fails with an error message:

" While running the child process: Child was terminated by signal 6 (SIGABRT)"

Error in cleaning up after application: Exception: Error reading fast program

output: Unable to open fastx output file: "/tmp/bslskAAAMGXMCf.tmp" (at

/tmp/bslskAAAMGXMCf.tmp:0)."

Workaround

There is an upper limit on the amount of memory that is allocated per process. For tru64 machine the limit for datasize is set to 128M. To increase this limit, execute

unlimit datasize (csh) or

ulimit datasize (ksh)

This will increase the limit on the datasize to 1024M. This is the maximum amount of memory that an individual process can take on Tru64 machine. So, default settings for the search set parameter (-infile2) for the fasta suite of programs may cause a crash. Please execute the programs with a smaller subset. The programs have been tested successfully using a search set of 400 thousand sequences

ALGORITHM

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For a description of the algorithm, see the FastA+ program documentation. FastX+ always uses an unrestricted Smith-Waterman algorithm for the final alignment, so this step may take a long time.

CONSIDERATIONS

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Accelrys GCG (GCG) version of FastX+ searches using both strands of nucleic acid queries unless you use -onestrand. Dr. Pearson's FASTX+ searches with one strand only.

The E() scores are affected by similarities in sequence composition between the query sequence and the search set sequence. Unrelated sequences may have "significant" scores because of composition bias.

If there is a database entry that overlaps your query in several places, but there are large gaps between the matching regions, only the best overlap appears in the alignment display.

There are two ways to control the size of the list of best scores. By default, scores are listed until a specific E() value is reached. You may set the value in response to the program prompt or by using -expect; otherwise the program uses 10.0 for protein searches, 2.0 for nucleic acid searches. (If you are running the program interactively, it will show no more than 40 scores initially, and ask if you want to see more scores if there are any more that are less than the E() value.)

If you use -listsize, the E() value is ignored, and the program will list the number of scores you requested.

You can control the number of alignments using -noalign and -align. The program behaves differently depending on whether it is being run noninteractively (in batch or with -Default on the command line) or interactively. In the noninteractive case, the program displays the number of alignments set by -align. (If this is not present, it shows 40 alignments or the number of scores that were listed, whichever is smaller.) If you run the program interactively, it displays the list of best scores, and then asks you how many alignments you want to see. (This prompt does not appear if you use -noalign or -align.)

Increasing Sensitivity By Adjusting Word Size

By default, FastX+ uses a word size of 2. If it finds few or no matches, especially if your query sequence is short, rerun the search using -wordsize=1 to increase the sensitivity. Note that this will dramatically increase the amount of CPU time required to run the program.

Adjusting Gap Creation, Gap Extension, and Frameshift Penalties

Unlike other GCG programs, FastX+ does not read the default gap creation, gap extension, and frameshift penalties from the scoring matrix file. It uses default penalties that were empirically determined to be appropriate for the BLOSUM50 scoring matrix. If you select a different scoring matrix with -matrix, you may need to change the gap penalties. The histogram display gives a qualitative view of the quality of fit between the actual distribution of scores and the expected distribution of scores. This information may indicate whether or not suitable gap creation and extension penalties were used for the search. When the histogram shows poor agreement between the actual distribution and the theoretical distribution, you might consider using -gapweight and/or -lengthweight to specify higher gap creation and extension penalties, respectively. For example, you might increase the gap creation penalty from 15 to 20 and the gap extension penalty from 2 to 4. You may also need to use -frameweight to adjust the frameshift penalty.

Differences in Applying Gap Extension Penalties

There are two different philosophies on how to penalize gaps in an alignment. One way is to penalize a gap by the gap creation penalty plus the extension penalty times the length of the gap (gapweight + (lengthweight x gap length)). The other way is to use the gap creation penalty plus the extension penalty times the gap length excluding the first residue in the gap (gapweight + (lengthweight x (gap length - 1)).

"Native" GCG programs, such as Framesearch and Bestfit, handle gap extension penalties the first way, while the FastA+-family programs use the second way. Therefore a value for -lengthweight that gives good results with one of the FastA+-family programs may not give equivalent results with a native GCG program, and vice versa.

Increasing Program Speed Using Multithreading

This program is multithreaded. It has the potential to run faster on a machine equipped with multiple processors because different parts of the analysis can be run in parallel on different processors. By default, the program assumes you have one processor, so the analysis is performed using one thread. You can use -processors to increase the number of threads up to the number of physical processors on the computer.

Under ideal conditions, the increase in speed is roughly linear with the number of processors used. But conditions are rarely ideal. If your computer is heavily used, competition for the processors can reduce the program's performance. In such an environment, try to run multithreaded programs during times when the load on the system is light.

As the number of threads increases, the amount of memory required increases substantially. You may need to ask your system administrator to increase the memory quota for your account if you want to use more than two threads.

Never use -processors to set the number of threads higher than the number of physical processors that the machine has -- it does not increase program performance, but instead uses up a lot of memory needlessly and makes it harder for other users on the system to get processor time. Ask your system administrator how many processors your computer has if you aren't sure.

SUGGESTIONS

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Identifying the Search Set

If you want to search a part of a database instead of an entire database, see the "Using Database Sequences" topic of Section 2, Using Sequence Files and Databases of the User's Guide for a list of the logical names used for the databases and the divisions of each database. The search set can also consist of a group of sequence files that are not in a database. Use a multiple sequence specification to name these. For information about naming groups of sequences for the search set, see the topics "Specifying Files" and "Using Wildcards" in Section 1, Getting Started, and "Using Database Sequences," "Using Multiple Sequence Format (MSF) Files", "Using Rich Sequence Format (RSF) Files", and "Using List Files" in Section 2, Using Sequence Files and Databases of the User's Guide.

Batch Queue

FastX+ is one of the few programs in GCG that can take more than a few minutes to run. Most comparisons should probably be run in the batch queue. You can specify that this program run at a later time in the batch queue by using -batch. Run this way, the program prompts you for all the required parameters and then automatically submits itself to the batch or at queue. For more information, see "Using the Batch Queue" in Section 3, Using Programs in the User's Guide. Very large comparisons may exceed the CPU limit set by some systems.

Interrupting a Search: <Ctrl>C

You can type <Ctrl>C to interrupt a search and see the results from the part of the search that has already been completed. Because the program is multithreaded, the search may not be interrupted immediately, but will continue until one of the threads finishes processing its data and returns for more data.

COMMAND-LINE SUMMARY

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All parameters for this program may be added to the command line. Use -check to view the summary below and to specify parameters before the program executes. In the syntax summary below, square brackets ([ and ]) enclose parameter values that are optional. For each program parameter, square brackets enclose the type of parameter value specified, the default parameter value, and shortened forms of the parameter name, aliases. Programs with a plus in the name use either the full parameter name or a specified alias. If “Type” is “Boolean”, then the presence of the parameter on the command line indicates a true condition. A false condition needs to be stated as, parameter=false.

Minimal Syntax: % fastx+ [-infile1=]value –Default.

Minimal Parameters (case-insensitive):

-infile1 [Type: List / Default: EMPTY / Aliases: infile in1 in]

Input file specification.

Prompted Parameters (case-insensitive):

-begin [Type: Integer / Default: '1' / Aliases: beg]

Starting point of the range of interest in the input sequence.

-end [Type: Integer / Default: '-1']

End point of the range of interest in the input sequence. A value of '-1' indicates that the range extends till the end of input sequence.

-infile2 [Type: List / Default: EMPTY / Aliases: in2 db]

Search set specification.

-outfile [Type: OutFile / Default: '<sequence_name>.<program_name>' /Aliases: out]File to which output is written. A value of '-' means STDOUT. Specifying this option also turns on the 'concat' option.Default value is '-'.

Optional Parameters (case-insensitive):

-check [Type: Boolean / Default: 'false' / Aliases: che help]

Prints out this usage message.

-default [Type: Boolean / Default: 'false' / Aliases: d def]

Specifies that sensible default values be used for all parameters where possible.

-documentation [Type: Boolean / Default: 'true' / Aliases: doc]

Prints banner at program startup.

-quiet [Type: Boolean / Default: 'false' / Aliases: qui]

Tells application to print only a minimal amount of information.

-wordsize [Type: Integer / Default: EMPTY / Aliases: wor]

Size of word (k-tuple) used in the hashing step.

-expect [Type: Double / Default: '2.0' / Aliases: exp]

Shows all scores whose E() value is less than the specified value of expect.

-matrix [Type: String / Default: EMPTY / Aliases: mat]

Assigns the scoring matrix for the comparison.

-processors [Type: Integer / Default: '1' / Aliases: proc]

On a multiprocessor computer, this parameter controls the number of threads to use for database search.

-minlength [Type: Integer / Default: EMPTY / Aliases: minl]

The search set is restricted to sequences whose length is more than the value specified by this parameter.

-maxlength [Type: Integer / Default: EMPTY / Aliases: maxl]

The search set is restricted to sequences whose length is less than the value specified by this parameter.

-pamfactor [Type: Boolean / Default: 'DEFAULT_PARAM_VALUE' / Aliases:pam] This parameter governs whether a scoring matrix should be used for calculating initial diagonal scores, instead of using the identical match scores from the scoring matrix.

Default is to use FASTA internal behavior, which differs for protein and nucleotide searches.

-gapweight [Type: Integer / Default: EMPTY / Aliases: gap]

This parameter specifies the gap creation penalty that is substracted from an alignment every time a gap is created.

-lengthweight [Type: Integer / Default: EMPTY / Aliases: len]

This parameter specifies the gap extension penalty that is substracted from an alignment every time a gap is extended by one residue.

-optall [Type: Boolean / Default: 'DEFAULT_PARAM_VALUE' / Aliases:opt]With this parameter, the program immediately performs an alignment and calculates the opt score when the initn score is greater than or equal to the value specified by this parameter. This parameter allows you to override the default threshold calculated by the program. Scores are sorted and saved by opt score during the search. -NOOPTall doesn't compute the opt score until the search is complete. In this case scores are sorted and saved by initn score instead of by opt score.

-listsize [Type: Integer / Default: '10' / Aliases: lis]

This parameter controls the number of top scores show. Overrides the -EXPect parameter.

-alignments [Type: Integer / Default: '20' / Aliases: align ali]

This parameter limits the number of alignments to display in the output file to the 10 best matches in the list. Use -NOALIgn to suppress the sequence alignments in the output file.

-showall [Type: Boolean / Default: 'DEFAULT_PARAM_VALUE' / Aliases:show]Shows entire sequences in the alignment display, instead of just the best region of overlap and its surroundings.

-native [Type: Boolean / Default: 'false']

Output native fasta formatted output.

-markx [Type: Integer / Default: EMPTY / Aliases: mark]

This parameter determines the alignment display mode - especially the symbols that identify matches and mismatches. The default value, -MARKx=0 uses a colon to show identities and a period (.) to show conservative replacements.

-MARKx=1 will not mark identities; instead, conservative replacements are connected with a lowercase x, and non-conservative substitutions are connected with an uppercase X.

If -MARKx=2, the residues in the second sequence are shown only if they differ from the first sequence.

-MARKx=3 displays the aligned library sequences without the query sequences; these can be used to build a primitive multiple alignment.

-MARKx=4 provides a graphical display of the boundaries of the alignments.

-MARKx=5 provides a combination of -MARKx=4 and -MARKx=0.

-MARKx=6 provides -MARKx=5 plus HTML formatting.

-MARKx=9 provides percent identity and coordinates with the initial list of high scores as well as the conventional

-MARKx=0 alignments.

Use -MARKx=10 to get aligned sequences in the FastA "parsable" output format.

-histogram [Type: Boolean / Default: 'true' / Aliases: his]

Start/Suppress printing the histogram.

-linesize [Type: Integer / Default: EMPTY / Aliases: lin]

This parameter lets you set the number of sequence symbols in each line of the alignment to any number between 60 and 200.

-batch [Type: Boolean / Default: 'false']

Allows submitting a job to a batch queue.

-onestrand [Type: Boolean / Default: 'false' / Aliases: one]

Translates only the 3 forward frames.

-frameweight [Type: Integer / Default: EMPTY / Aliases: frame fra]

LOCAL DATA FILES

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The files described below supply auxiliary data to this program. The program automatically reads them from a public data directory unless you either 1) have a data file with exactly the same name in your current working directory; or 2) name a file on the command line with an expression like -DATa1=myfile.dat. For more information see Section 4, Using Data Files in the User's Guide.

Local Scoring Matrices

This program reads one or more scoring matrices for the comparison of sequence characters. The program automatically reads the program's default scoring matrix in a public data directory unless you either

1) Have a data file with exactly the same name as the program default scoring matrix in your current working directory; or

2) Have a data file with exactly the same name as the program default scoring matrix in the directory with the logical name Share_Matrix; or

3) Name a file on the command line with an expression like -matrix=mymatrix.cmp. If you don't include a directory specification when you name a file with -matrix, the program searches for the file first in your local directory, then in the directory with the logical name Share_Matrix,. For more information see "Using a Special Kind of Data File: A Scoring Matrix" in Section 4, Using Data Files in the User's Guide.

FastX+ reads a scoring matrix containing the values for every possible match from your working directory or the public database. The file blosum50.cmp contains the default values for matches. blosum50.cmp is a BLOSUM50 matrix. You can use the Fetch+ program to obtain a copy of this file in order to modify it to suit your own needs.

PARAMETER REFERENCE

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You can set the parameters listed below from the command line. Shortened forms of the parameter name, aliases, are shown, separated by commas.

-infile1, infile, -in1, -in

Inputs file specification.

-begin, -beg

Starting point of the range of interest in the input sequence.

-end

End point of the range of interest in the input sequence. A value of '-1' indicates that the range extends till the end of input sequence.

-infile2, -in2, -db

Search set specification.

-outfile, -out

File to which output is written. A value of '-' means STDOUT. Specifying this option also turns on the 'concat' option. Default value is '-'

-wordsize=2, -wor

Sets the size of the word (k-tuple) to use for the hashing step.

-matrix=mymatrix.cmp, -mat

Allows you to specify a scoring matrix file name other than the program default. If you don't include a directory specification when you name a file with -matrix, the program searches for the file first in your local directory, then in the directory with the logical name MyData, then in the public data directory with the logical name GenMoreData, and finally in the public data directory with the logical name GenRunData.

For more information see the Local Scoring Matrices section.

-expect=2.0, -exp

Shows all scores whose E() value is less than 2.0. Ignored if -listsize is used.

-check, -che, -help

Prints out this usage message.

-default, -d, -def

Specifies that sensible default values be used for all parameters where possible.

-documentation, -doc

Prints banner at program startup.

-quiet, -qui

This parameter is not supported.

-alignments, -align, -ali

This parameter limits the number of alignments to display in the output file to the 10 best matches in the list. Use -noalignn to suppress the sequence alignments in the output file.

-histogram, -his

Start/suppress printing the histogram.

-markx, -mark

This parameter determines the alignment display mode - especially the symbols that identify matches and mismatches. The default value, -markx=0 uses a colon to show identities and a period (.) to show conservative replacements.

-markx=1 will not mark identities; instead, conservative replacements are connected with a lowercase x, and non-conservative substitutions are connected with an uppercase X.

If -markx=2, the residues in the second sequence are shown only if they differ from the first sequence.

-markx=3 displays the aligned library sequences without the query sequences; these can be used to build a primitive multiple alignment.

-markx=4 provides a graphical display of the boundaries of the alignments.

-markx=5 provides a combination of -markx=4 and -markx=0.

-markx=6 provides -markx=5 plus HTML formatting.

-markx=9 provides percent identity and coordinates with the initial list of high scores as well as the conventional

-markx=0 alignments.

Use -markx=10 to get aligned sequences in the FastA "parsable" output format.

-linesize=60, -lin

Lets you set the number of sequence symbols in each line of the alignment to any number between 60 and 200.

-processors=2, -proc

Tells the program to use 2 threads for the database search on a multiprocessor computer.

-minlength=1000, -minl

Restricts the search to search set sequences that are equal to or longer than 1000 residues.

-maxlength=5000, -maxl

Restricts the search to search set sequences that are equal to or shorter than 5000 residues.

-onestrand, -one

Searches using only the top strand of the query sequence.

-pamfactor, -pam

Uses a scoring matrix for the calculation of initial diagonal scores. Instead of using a constant factor for each match in a word, values are obtained from a scoring matrix. This is the default option for protein sequences.

-gapweight=12, -gap

Specifies the gap creation penalty that is subtracted from the alignment score whenever a gap is created.

-lengthweight=2, -len

Specifies the gap extension penalty that is subtracted from the alignment score for each residue added to an existing gap.

-frameweight=20, -frame

Specifies the penalty that is subtracted from the alignment score whenever a frameshift is inserted.

-optall=20, -opt

Immediately performs an alignment and calculates the opt score when the initn score is greater than or equal to 20. This parameter allows you to override the default threshold calculated by the program. Scores are sorted and saved by opt score during the search. -nooptall doesn't compute the opt score until the search is complete. In this case scores are sorted and saved by initn score instead of by opt score.

-native

Output native fasta formatted output.

-listsize=40, -lis

Shows the best 40 scores. Overrides -expect.

-showall, -show

Shows entire sequences in the alignment display, instead of just the best region of overlap and its surroundings.

-linesize=60, -lin

Lets you set the number of sequence symbols in each line of the alignment to any number between 60 and 200.

-batch, -bat

Submits the program to the batch queue for processing after prompting you for all required user inputs. Any information that would normally appear on the screen while the program is running is written into a log file. Whether that log file is deleted, printed, or saved to your current directory depends on how your system manager has set up the command that submits this program to the batch queue. All output files are written to your current directory, unless you direct the output to another directory when you specify the output file.

Printed: September 9, 2005 16:19

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