est2genome.txt

emboss的linux版本的源代码

                                est2genome 



Function

   Align EST and genomic DNA sequences

Description

   est2genome is a software tool to aid the prediction of genes by
   sequence homology. The program will align a set of spliced nucleotide
   sequences (ESTs cDNAs or mRNAs) to an unspliced genomic DNA sequence,
   inserting introns of arbitrary length when needed. In addition, where
   feasible introns start and stop at the splice consensus dinucleotides
   GT and AG.

   Unless instructed otherwise, the program makes three alignments: First
   it compares both stands of the spliced sequence against the forward
   strand of the genomic, assuming the splice consensus GT/AG (ie in the
   forward gene direction). The maximum-scoring orientation is then
   realigned assuming the splice consensus CT/AC (ie in the reversed gene
   direction). Only the overall maximum-scoring alignment is reported.

   The program outputs a list of the exons and introns it has found. The
   format is like that of MSPcrunch, ie a list of matching segments. This
   format is easy to parse into other software. The program also
   indicates, based on the splice site information, the gene's predicted
   direction of transcription. Optionally the full sequence alignment is
   printed as well (see the example).

Algorithm

   The program uses a linear-space divide-and-conquer strategy (Myers and
   Miller, 1988; Huang, 1994) to limit memory use:

   1. A first pass Smith-Waterman local alignment scan is done to find
   the start and end of the maximally scoring segments.

   2. Subsequences corresponding to these segments are extracted

   3a. If the product of the subsequences' lengths is less than a
   user-defined threshold (i.e. they will fit in memory) the segments are
   realigned using the Needleman-Wunsch global alignment algorithm, which
   will give the same result as the Smith-Waterman since the subsequences
   are guaranteed to align end-to-end.

   3b. If the product of the lengths exceeds the threshold (a full
   alignment will not fit in memory) the alignment is made recursively by
   splitting the spliced (EST) sequence in half and finding the genome
   sequence position which aligns with the mid-point. The process is
   repeated until the product of gthe lengths is less than the threshold.
   The divided sequences are aligned separately and then merged.

   4. The genome sequence is searched against the forward and reverse
   strands of the spliced (EST) sequence, assuming a forward gene
   splicing direction (i.e. GT/AG consensus).

   5. Then the best-scoring orientation is realigned assuming reverse
   splicing (CT/AC consensus). The overall best alignment is reported.

Usage

   Here is a sample session with est2genome


% est2genome 
Align EST and genomic DNA sequences
Spliced EST nucleotide sequence(s): tembl:hs989235
Unspliced genomic nucleotide sequence: tembl:hsnfg9
Output file [hs989235.est2genome]: 

   Go to the input files for this example
   Go to the output files for this example

Command line arguments

   Standard (Mandatory) qualifiers:
  [-estsequence]       seqall     Spliced EST nucleotide sequence(s)
  [-genomesequence]    sequence   Unspliced genomic nucleotide sequence
  [-outfile]           outfile    [*.est2genome] Output file name

   Additional (Optional) qualifiers:
   -match              integer    [1] Score for matching two bases (Any
                                  integer value)
   -mismatch           integer    [1] Cost for mismatching two bases (Any
                                  integer value)
   -gappenalty         integer    [2] Cost for deleting a single base in
                                  either sequence, excluding introns (Any
                                  integer value)
   -intronpenalty      integer    [40] Cost for an intron, independent of
                                  length. (Any integer value)
   -splicepenalty      integer    [20] Cost for an intron, independent of
                                  length and starting/ending on donor-acceptor
                                  sites (Any integer value)
   -minscore           integer    [30] Exclude alignments with scores below
                                  this threshold score. (Any integer value)

   Advanced (Unprompted) qualifiers:
   -reverse            boolean    Reverse the orientation of the EST sequence
   -[no]splice         boolean    [Y] Use donor and acceptor splice sites. If
                                  you want to ignore donor-acceptor sites then
                                  set this to be false.
   -mode               menu       [both] This determines the comparion mode.
                                  The default value is 'both', in which case
                                  both strands of the est are compared
                                  assuming a forward gene direction (ie GT/AG
                                  splice sites), and the best comparsion
                                  redone assuming a reversed (CT/AC) gene
                                  splicing direction. The other allowed modes
                                  are 'forward', when just the forward strand
                                  is searched, and 'reverse', ditto for the
                                  reverse strand. (Values: both (Both
                                  strands); forward (Forward strand only);
                                  reverse (Reverse strand only))
   -[no]best           boolean    [Y] You can print out all comparisons
                                  instead of just the best one by setting this
                                  to be false.
   -space              float      [10.0] For linear-space recursion. If
                                  product of sequence lengths divided by 4
                                  exceeds this then a divide-and-conquer
                                  strategy is used to control the memory
                                  requirements. In this way very long
                                  sequences can be aligned.
                                  If you have a machine with plenty of memory
                                  you can raise this parameter (but do not
                                  exceed the machine's physical RAM) (Any
                                  numeric value)
   -shuffle            integer    [0] Shuffle (Any integer value)
   -seed               integer    [20825] Random number seed (Any integer
                                  value)
   -align              boolean    Show the alignment. The alignment includes
                                  the first and last 5 bases of each intron,
                                  together with the intron width. The
                                  direction of splicing is indicated by angle
                                  brackets (forward or reverse) or ????
                                  (unknown).
   -width              integer    [50] Alignment width (Any integer value)

   Associated qualifiers:

   "-estsequence" associated qualifiers
   -sbegin1            integer    Start of each sequence to be used
   -send1              integer    End of each sequence to be used
   -sreverse1          boolean    Reverse (if DNA)
   -sask1              boolean    Ask for begin/end/reverse
   -snucleotide1       boolean    Sequence is nucleotide
   -sprotein1          boolean    Sequence is protein
   -slower1            boolean    Make lower case
   -supper1            boolean    Make upper case
   -sformat1           string     Input sequence format
   -sdbname1           string     Database name
   -sid1               string     Entryname
   -ufo1               string     UFO features
   -fformat1           string     Features format
   -fopenfile1         string     Features file name

   "-genomesequence" associated qualifiers
   -sbegin2            integer    Start of the sequence to be used
   -send2              integer    End of the sequence to be used
   -sreverse2          boolean    Reverse (if DNA)
   -sask2              boolean    Ask for begin/end/reverse
   -snucleotide2       boolean    Sequence is nucleotide
   -sprotein2          boolean    Sequence is protein
   -slower2            boolean    Make lower case
   -supper2            boolean    Make upper case
   -sformat2           string     Input sequence format
   -sdbname2           string     Database name
   -sid2               string     Entryname
   -ufo2               string     UFO features
   -fformat2           string     Features format
   -fopenfile2         string     Features file name

   "-outfile" associated qualifiers
   -odirectory3        string     Output directory

   General qualifiers:
   -auto               boolean    Turn off prompts
   -stdout             boolean    Write standard output
   -filter             boolean    Read standard input, write standard output
   -options            boolean    Prompt for standard and additional values
   -debug              boolean    Write debug output to program.dbg
   -verbose            boolean    Report some/full command line options
   -help               boolean    Report command line options. More
                                  information on associated and general
                                  qualifiers can be found with -help -verbose
   -warning            boolean    Report warnings
   -error              boolean    Report errors
   -fatal              boolean    Report fatal errors
   -die                boolean    Report dying program messages

Input file format

   est2genome reads two nucleotide sequences. The first is an EST
   sequence (a single read or a finished cDNA). The second is a genomic
   finished sequence.

  Input files for usage example

   'tembl:hs989235' is a sequence entry in the example nucleic acid
   database 'tembl'

  Database entry: tembl:hs989235

ID   HS989235   standard; RNA; EST; 495 BP.
XX
AC   H45989;
XX
SV   H45989.1
XX
DT   18-NOV-1995 (Rel. 45, Created)
DT   04-MAR-2000 (Rel. 63, Last updated, Version 2)
XX
DE   yo13c02.s1 Soares adult brain N2b5HB55Y Homo sapiens cDNA clone
DE   IMAGE:177794 3', mRNA sequence.
XX
KW   EST.
XX
OS   Homo sapiens (human)
OC   Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia
;
OC   Eutheria; Primates; Catarrhini; Hominidae; Homo.
XX
RN   [1]
RP   1-495
RA   Hillier L., Clark N., Dubuque T., Elliston K., Hawkins M., Holman M.,
RA   Hultman M., Kucaba T., Le M., Lennon G., Marra M., Parsons J., Rifkin L.,
RA   Rohlfing T., Soares M., Tan F., Trevaskis E., Waterston R., Williamson A.,
RA   Wohldmann P., Wilson R.;
RT   "The WashU-Merck EST Project";
RL   Unpublished.
XX
DR   RZPD; IMAGp998F03326; IMAGp998F03326.
XX
CC   On May 8, 1995 this sequence version replaced gi:800819.
CC   Contact: Wilson RK
CC   Washington University School of Medicine
CC   4444 Forest Park Parkway, Box 8501, St. Louis, MO 63108
CC   Tel: [phone]
CC   Fax: [phone]
CC   Email: [e-mail]
CC   Insert Size: 544
CC   High quality sequence stops: 265
CC   Source: IMAGE Consortium, LLNL
CC   This clone is available royalty-free through LLNL ; contact the
CC   IMAGE Consortium ([e-mail]) for further information.
CC   Possible reversed clone: polyT not found
CC   Insert Length: 544   Std Error: 0.00
CC   Seq primer: SP6
CC   High quality sequence stop: 265.
XX
FH   Key             Location/Qualifiers
FH
FT   source          1..495
FT                   /db_xref="taxon:9606"
FT                   /db_xref="ESTLIB:300"
FT                   /db_xref="RZPD:IMAGp998F03326"
FT                   /note="Organ: brain; Vector: pT7T3D (Pharmacia) with a
FT                   modified polylinker; Site_1: Not I; Site_2: Eco RI; 1st
FT                   strand cDNA was primed with a Not I - oligo(dT) primer [5'
FT                   TGTTACCAATCTGAAGTGGGAGCGGCCGCGCTTTTTTTTTTTTTTTTTTT 3'],
FT                   double-stranded cDNA was size selected, ligated to Eco RI
FT                   adapters (Pharmacia), digested with Not I and cloned into
FT                   the Not I and Eco RI sites of a modified pT7T3 vector
FT                   (Pharmacia). Library went through one round of
FT                   normalization to a Cot = 53. Library constructed by Bento
FT                   Soares and M.Fatima Bonaldo. The adult brain RNA was
FT                   provided by Dr. Donald H. Gilden. Tissue was acquired 17-1
8
FT                   hours after death which occurred in consequence of a
FT                   ruptured aortic aneurysm. RNA was prepared from a pool of
FT                   tissues representing the following areas of the brain:
FT                   frontal, parietal, temporal and occipital cortex from the
FT                   left and right hemispheres, subcortical white matter, basa
l
FT                   ganglia, thalamus, cerebellum, midbrain, pons and medulla.
"
FT                   /sex="Male"
FT                   /organism="Homo sapiens"
FT                   /clone="IMAGE:177794"
FT                   /clone_lib="Soares adult brain N2b5HB55Y"
FT                   /dev_stage="55-year old"
FT                   /lab_host="DH10B (ampicillin resistant)"
XX
SQ   Sequence 495 BP; 73 A; 135 C; 169 G; 104 T; 14 other;
     ccggnaagct cancttggac caccgactct cgantgnntc gccgcgggag ccggntggan        6
0
     aacctgagcg ggactggnag aaggagcaga gggaggcagc acccggcgtg acggnagtgt       12
0
     gtggggcact caggccttcc gcagtgtcat ctgccacacg gaaggcacgg ccacgggcag       18
0
     gggggtctat gatcttctgc atgcccagct ggcatggccc cacgtagagt ggnntggcgt       24
0
     ctcggtgctg gtcagcgaca cgttgtcctg gctgggcagg tccagctccc ggaggacctg       30
0
     gggcttcagc ttcccgtagc gctggctgca gtgacggatg ctcttgcgct gccatttctg       36
0
     ggtgctgtca ctgtccttgc tcactccaaa ccagttcggc ggtccccctg cggatggtct       42
0
     gtgttgatgg acgtttgggc tttgcagcac cggccgccga gttcatggtn gggtnaagag       48
0
     atttgggttt tttcn                                                        49
5
//

  Database entry: tembl:hsnfg9

ID   HSNFG9     standard; DNA; HUM; 33760 BP.
XX
AC   Z69719;
XX
SV   Z69719.1
XX
DT   26-FEB-1996 (Rel. 46, Created)
DT   22-NOV-1999 (Rel. 61, Last updated, Version 3)
XX
DE   Human DNA sequence from cosmid NFG9 from a contig from the tip of the shor
t
DE   arm of chromosome 16, spanning 2Mb of 16p13.3. Contains Interleukin 9
DE   Receptor Pseudogene, repeat polymorphism, ESTs, CpG islands and endogenous
DE   retroviral DNA.
XX
KW   16p13.3; CpG island; Interleukin 9 Receptor Pseudogene;
KW   repeat polymorphism.
XX
OS   Homo sapiens (human)
OC   Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia
;
OC   Eutheria; Primates; Catarrhini; Hominidae; Homo.
XX
RN   [1]
RP   1-33760
RA   Kershaw J.;
RT   ;
RL   Submitted (22-FEB-1996) to the EMBL/GenBank/DDBJ databases.
RL   Sanger Centre, Hinxton, Cambridgeshire, CB10 1RQ, England. E-mail enquires
:
RL   [e-mail]