2244 lines
94 KiB
Text
2244 lines
94 KiB
Text
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@-1. TX 0 @General
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@-2. T 0 @Screen control
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@-2. X 0 @Screen
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@-3. T 0 @Statistical analysis of content
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@-3. X 0 @Statistics
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@-4. T 0 @Structures and repeats
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@-4. X 0 @Structures
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@-5. TX 0 @Search
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@0. TX -1 @PIP
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This is a program for analysing individual protein sequences.
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It can read sequences stored in many of the most commonly used
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formats, and performs all of the usual simple analyses. In addition
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it has very flexible search procedures and presents many of its
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results graphically.
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The following analyses (preceded by their option numbers) are
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included:
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? = Help
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! = Quit
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3 = read a new sequence
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4 = define active region
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5 = list the sequence
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6 = list a text file
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7 = direct output to disk
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8 = write active sequence to disk
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9 = edit the sequence
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10 = clear graphics screen
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11 = clear text screen
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12 = draw a ruler
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13 = use cross hair
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14 = reposition plots
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15 = label diagram
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16 = display a map
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17 = search for short sequences
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18 = compare a sequence
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19 = compare a sequence using a score matrix
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20 = search for a sequence using a weight matrix
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21 = calculate amino acid composition
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22 = plot hydrophobicity
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23 = plot charge
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24 = plot Robson prediction
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25 = plot hydrophobic moment
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26 = draw helix wheel
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27 = back translate
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28 = search for patterns of motifs
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Some of these methods produce graphical results and so the
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program is generally used from a graphics terminal (a vdu on which
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lines and points can be drawn as well as characters).
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For users of VT640's or their equivalents the terminal must be
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set nowrap (type NOWRAP) prior to running the program.
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The positions of each of the plots is defined relative to a users
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drawing board which has size 1-10,000 in x and 1-10,000 in y. Plots
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for each option are drawn in a window defined by x0,y0 and
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xlength,ylength. Where x0,y0 is the position of the bottom left hand
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corner of the window, and xlength is the width of the window and
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ylength the height of the window.
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--------------------------------------------------------- 10,000
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1 1
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1 -------------------------------------- ^ 1
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1 1 1 1 1
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1 1 1 1 1
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1 1 1 ylength 1
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1 1 1 1 1
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1 1 1 1 1
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1 -------------------------------------- v 1
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1 x0,y0^ 1
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1 <---------------xlength--------------> 1
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--------------------------------------------------------- 1
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1 10,000
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All values are in drawing board units (i.e. 1-10,000, 1-10,000).
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The default window positions are read from a file "ANALPMRG" when
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the program is started. Users can have their own file if required.
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The program can handle sequences stored in several formats:
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Staden, EMBL, GENBANK, PIR (also known as NBRF) and GCG and they are
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described in the help for 'READ NEW SEQUENCE'.
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The options for the program are accessed from 5 main menus:
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general, screen control, statistical analysis of content, structure,
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search. Both menus and options are selected by number.
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@1. TX 0 @Help
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This option gives online help. The user should select option
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numbers and the current documentation will be given. Note that
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option 0 gives an introduction to the program, and that ? will get
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help from anywhere in the program. The following analyses (preceded
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by their option numbers) are included:
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@2. TX 0 @Quit
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This function stops the program.
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@3. TX 1 @Read a new sequence
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This option allows users to read in new sequences, browse
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through annotations, or search sequence libraries for keywords.
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Sequences can be read from "personal" sequence files or from
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sequence libraries. These are referred to as the sequence "source".
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Personal files can be stored in several formats: Staden, PIR, EMBL,
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GENBANK and GCG. At LMB we use "Staden" format for sequencing and
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all the libraries are stored in their original formats. Note,
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however, that libraries such as EMBL or GenBank that are divided
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into several files (eg GenBank has 13 separate files) are indexed as
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a whole. This means that users do not need to know which file
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contains an entry, only which library. When the user selects to
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read in a sequence the program first asks for the sequence "source".
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If the user selects "personal" the program will ask for the
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format (Staden, PIR, EMBL, GENBANK or GCG), and then for the name of
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the file. For PIR format the user will also be required to know the
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entry name of the sequence as the file can contain several. For the
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other formats only a single entry is expected. The file will be
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read, its length and composition will be displayed and the option
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left.
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If the user selects "library" as the sequence source the
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program will display a list of available libraries. The programs are
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capable of handling all current libraries but which ones are
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available will vary from site to site. At LMB we have several
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libraries and also weekly updates of data gathered between releases.
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The program will ask users to select a library and then give a list
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of options:
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X 1 Get a sequence
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2 Get annotations
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3 Get entrynames from accession numbers
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4 Search titles for keywords
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5 Search text index for keywords
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If get a sequence or get annotations is selected users will be asked
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to type the entry name. The option will be left when a sequence is
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selected or ! is typed. The composition and length will be
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displayed.
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The text index contains all words from feature tables,
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reference titles, definition lines, keywords lists and comments, so
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the text index search is most useful. It is also the fastest. Up to
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5 words can be searched for at once. The words should be typed
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separated by spaces, for example
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? Keywords=P53 mouse murine tumo
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will search for all entries that contain words starting with p53,
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mouse, murine and tumo. Only the unique entries that contain ALL
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words will be listed. Before listing the matching entries the
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program will show the number of 'hits' for each word and ring the
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bell. Escape is possible at this point, or after each screenfull of
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entries. In addition to the entry names the text search displays
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the primary accession number, the sequence length and up to 80
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characters of description. (The search of 'titles' is now redundant
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because the full text index contains all the title words and the
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search is much faster. It will probably be removed from the
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program.) All searches are independent of case. Where possible the
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program will offer default entry names.
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Typical dialogue follows.
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Select sequence source
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X 1 Personal file
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2 Sequence library
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? Selection (1-2) (1) =
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Select sequence file format
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X 1 Staden
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2 EMBL
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3 GenBank
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4 PIR
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5 GCG
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? Selection (1-5) (1) =
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? Sequence file name=M13MP7.SEQ
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Contig title removed
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Sequence length= 7238
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Sequence composition
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T C A G -
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2405. 1539. 1765. 1527. 2.
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33.2% 21.3% 24.4% 21.1% 0.0%
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.
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.
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Select sequence source
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X 1 Personal file
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2 Sequence library
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? Selection (1-2) (1) =2
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Select a library
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X 1 EMBL 29 nucleotide library Dec 91
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2 SWISSPROT 20 protein library Nov 91
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3 PIR 31 protein library Dec 91
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4 NRL3D 58 From Brookhaven protein library Dec 91
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5 GenBank
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? Selection (1-5) (1) =
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Library is in EMBL format with indexes
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Select a task
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X 1 Get a sequence
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2 Get annotations
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3 Get entry names from accession numbers
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4 Search titles for keywords
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5 Search text index for keywords
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? Selection (1-5) (1) =5
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Search for keywords
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? Keywords=P53 mouse
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P53 hits 68
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MOUSE hits 8180
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MMANT01 X00875 536 Murine gene fragment for cellular tumour antigen
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MMANT02 X00876 83 Murine gene fragment for cellular tumour antigen
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MMANT03 X00877 21 Murine gene fragment for cellular tumour antigen
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MMANT04 X00878 261 Murine gene fragment for cellular tumour antigen
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MMANT05 X00879 184 Murine gene fragment for cellular tumour antigen
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MMANT06 X00880 113 Murine gene fragment for cellular tumour antigen
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MMANT07 X00881 110 Murine gene fragment for cellular tumour antigen
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MMANT08 X00882 137 Murine gene fragment for cellular tumour antigen
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MMANT09 X00883 74 Murine gene fragment for cellular tumour antigen
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MMANT10 X00884 107 Murine gene for cellular tumour antigen p53 (exon
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MMANT11 X00885 562 Murine p53 gene 3' region with exon 11
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MMANTP53 M26862 536 Mouse tumor antigen p53 gene, 5' end.
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MMLYN M64608 2044 Mouse lyn protein mRNA, complete cds.
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MMP53 X00741 1377 Mouse mRNA for transformation associated protein
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MMP53A M13872 1285 Mouse p53 mRNA, complete cds, clone pcD53.
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MMP53B M13873 1241 Mouse p53 mRNA, complete cds, clone p53-m11.
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MMP53C M13874 1322 Mouse p53 mRNA, complete cds, clone p53-m8.
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MMP53G1 X01235 554 Mouse genomic DNA for 5' region of cellular tumou
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MMP53IN4 X60470 729 M.musculus p53 gene for p53 protein, intron 4
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MMP53P X01236 2132 Mouse pseudogene for cellular tumour antigen p53
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MMP53R X01237 1773 Mouse mRNA for cellular tumour antigen p53
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MMRSB2P5 M64597 196 Mouse B2 repeat in the 3' flank of protein 53 (p5
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22 different entries found
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Select a task
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X 1 Get a sequence
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2 Get annotations
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3 Get entry names from accession numbers
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4 Search titles for keywords
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5 Search text index for keywords
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? Selection (1-5) (1) =4
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Search for keywords
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? Keywords=alpha
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Searching for alpha
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AAGHA 623 a.anguilla mrna for glycoprotein hormone alpha subunit precu
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AAMALI 3338 a.aegypti mali gene encoding alpha 1-4 glucosidase, complete
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AAMALIA 1659 a.aegypti maltase-like i (mali) gene encoding alpha-1,4-gluc
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AAMALIB 1832 a.aegypti maltase-like i (mali) mrna encoding alpha-1,4-gluc
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ACA13GT 371 alouatta caraya alpha-1,3gt gene, 3' flank.
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ADHBADA1 102 duck alpha-d-globin gene, exon 1.
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ADHBADA2 1145 duck alpha-a-globin gene and 5' flank
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ADHBADWP 513 duck (white pekin) alpha ii (minor) globin mrna, complete co
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AEACOXABC 5279 a.eutrophus protein x (acox), acetoin:dcpip oxidoreductase-a
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AGA13GT 371 ateles geoffroyi alpha-1,3gt gene, 3' flank.
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AGAAAGFP 282 c.tetragonoloba alpha-amylase/alpha-galactosidase fusion pro
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AGAABL 138 b.subtilis alpha-amylase signal peptide gene e.coli beta-lac
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AGAFAMYA 57 synthetic b.stearothermophilus alpha amylase/s.cerevisiae ma
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AGAFAMYB 57 synthetic b.stearothermophilus alpha amylase/s.cerevisiae ma
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AGAFAMYC 57 synthetic b.stearothermophilus alpha amylase/s.cerevisiae ma
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AGAFCOXA 98 synthetic alpha-factor/cox iv fusion gene signal peptide.
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AGAGABA 7876 synthetic gossypium hirsutum (cotton) alpha globulin a and b
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AGAMYLS 120 synthetic alpha-amylase gene, 5' end.
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AGANPS 95 synthetic gene (jcnf-1) encoding alpha-factor pro-region/han
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!
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Select a task
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X 1 Get a sequence
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2 Get annotations
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3 Get entry names from accession numbers
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4 Search titles for keywords
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5 Search text index for keywords
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? Selection (1-5) (1) =3
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? Accession number=v00636
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Entry name LAMBDA
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Select a task
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X 1 Get a sequence
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2 Get annotations
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3 Get entry names from accession numbers
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4 Search titles for keywords
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5 Search text index for keywords
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? Selection (1-5) (1) =2
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Default Entry name=LAMBDA
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? Entry name=
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ID LAMBDA standard; DNA; PHG; 48502 BP.
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XX
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AC V00636; J02459; M17233; X00906;
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XX
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DT 03-JUL-1991 (Rel. 28, Last updated, Version 3)
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DT 09-JUN-1982 (Rel. 1, Created)
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XX
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DE Genome of the bacteriophage lambda (Styloviridae).
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XX
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KW circular; coat protein; DNA binding protein; genome;
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KW origin of replication.
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XX
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OS Bacteriophage lambda
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OC Viridae; ds-DNA nonenveloped viruses; Siphoviridae.
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XX
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RN [1]
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RP 1-48502
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RA Sanger F., Coulson A.R., Hong G.F., Hill D.F., Petersen G.B.;
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RT "Nucleotide sequence of bacteriophage lambda DNA";
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RL J. Mol. Biol. 162:729-773(1982).
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XX
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!
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Select a task
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X 1 Get a sequence
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2 Get annotations
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3 Get entry names from accession numbers
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4 Search titles for keywords
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5 Search text index for keywords
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? Selection (1-5) (1) =
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Default Entry name=LAMBDA
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? Entry name=
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DE Genome of the bacteriophage lambda (Styloviridae).
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Sequence length 48502
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Sequence composition
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T C A G -
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11988. 11360. 12336. 12818. 0.
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24.7% 23.4% 25.4% 26.4% 0.0%
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@4. TX 1 @Redefine active region
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For its analytic functions the program always works on a
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region of the sequence called the active region. When a new sequence
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is read into the program the active region is automatically set to
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start at the beginning of the sequence and go up to the maximum
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allowed size of active region the version of the program can handle.
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The positions are shown on the screen. On most machines this will
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be to the end of the sequence. This option allows the user define a
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different region. Note that for convenience in the listing and
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translation functions the user is given access to regions outside
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the active region.
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@5. TX 1 @List a sequence
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The sequence can be listed with line lengths from 10 to 120 in
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multiples of 10. Output can be directed to a disk file by first
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selecting disk output. The output looks like:
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10 20 30 40 50 60
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MQLNSTEISE LIKQRIAQFN VVSEAHNEGT IVSVSDGVIR IHGLADCMQG EMISLPGNRY
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70 80 90 100 110 120
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AIALNLERDS VGAVVMGPYA DLAEGMKVKC TGRILEVPVG RGLLGRVVNT LGAPIDGKGP
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130 140 150 160 170 180
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LDHDGFSAVE AIAPGVIERQ SVDQPVQTGY KAVDSMIPIG RGQRELIIGD RQTGKTALAI
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190 200 210 220 230 240
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DAIINQRDSG IKCIYVAIGQ KASTISNVVR KLEEHGALAN TIVVVATASE SAALQYLARM
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250 260 270 280 290 300
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PVALMGEYFR DRGEDALIIY DDLSKQAVAY RQISLLLRRP PGREAFPGDV FYLHSRLLER
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310 320 330 340 350 360
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AARVNAEYVE AFTKGEVKGK TGSLTALPII ETQAGDVSAF VPTNVISITD GQIFLETNLF
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370 380 390 400 410 420
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NAGIRPAVNP GISVSRVGGA AQTKIMKKLS GGIRTALAQY RELAAFSQFA SDLDDATRKQ
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430 440 450 460 470 480
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LDHGQKVTEL LKQKQYAPMS VAQQSLVLFA AERGYLADVE LSKIGSFEAA LLAYVDRDHA
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490 500 510 520 530 540
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PLMQEINQTG GYNDEIEGKL KGILDSFKAT QSW*
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@6. TX 1 @List a text file
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Allows the user to have a text file displayed on the screen.
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It will appear one page at a time.
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@7. TX 1 @Direct output to disk
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Used to direct output that would normally appear on the screen
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to a file.
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Select redirection of either text or graphics, and supply the
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name of the file that the output should be written to.
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The results from the next options selected will not appear on
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the screen but will be written to the file. When option 7 is
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selected again the file will be closed and output will again appear
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on the screen.
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@8. TX 1 @Write active region to disk
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The program has the capability of reading in EMBL, GENBANK,
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NBRF, GCG and Staden formats and of reversing and complementing
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sequences. This option allows users to write the current active
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sequence to a disk file in Staden format. Hence it allows format
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conversion and crude sequence cutting.
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@9. TX 1 @Edit the sequence
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Used to edit sequences or any other files by giving access to
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the computers system editor. For editing sequences the input file
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should have already been created using the listing function "list
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sequence".
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Supply the name of the file to edit. Wait while the system
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editor is made ready (can take awhile on a vax). Use the editor.
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Exit from the editor. If a sequence has been edited, and you want to
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process it, affirm that the sequence should be "made active". The
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edited sequence will replace the original sequence.
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This editing method is designed to give users access to an
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editor with which they are familiar - i.e. the one on their machine,
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and yet to allow them to edit a sequence which contains the
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landmarks they need in order to know where they are. Users can
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create files containing simple listings with numbering, using "list
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the sequence", and then edit them with their system editor, using
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the numbering to know where they are within the sequence. When the
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edits are complete they exit from the editor and the program
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"analyses" the edited file to extract only the sequence characters.
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Define the permitted set of characters to be:
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ACDEFGHIKLMNPQRSTVWXYZ-acdefghiklmnpqrstvwxyz. All permitted
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characters found in the file will become part of the sequence, all
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others removed.
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@10. TX 2 @Clear graphics
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Clears the screen of both text and graphics.
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@11. TX 2 @Clear text
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Clears only text from the screen.
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@12. TX 2 @Draw a ruler
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This option allows the user to draw a ruler or scale along the
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x axis of the screen to help identify the coordinates of points of
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interest. The user can define the position of the first amino acid
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to be marked (for example if the active region is 1501 to 8000, the
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user might wish to mark every 1000th amino acid starting at either
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1501 or 2000 - it depends if the user wishes to treat the active
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region as an independent unit with its own numbering starting at its
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left edge, or as part of the whole sequence). The user can also
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define the separation of the ticks on the scale and their height. If
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required the labelling routine can be used to add numbers to the
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ticks.
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@13. TX 2 @Use cross hair
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This function puts a steerable cross on the screen that can be
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used to find the coordinates of points in the sequence. The user can
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move the cross around using the directional keys; when he hits the
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space bar the program will print out the coordinates of the cross in
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sequence units and the option will be exited.
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If instead, you hit a , the position will be displayed but the
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cross will remain on the screen.
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If a letter s is hit the sequence around the cross hair is
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displayed and the cross remains on the screen.
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@14. TX 2 @Reset margins
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The positions of each of the plots is defined relative to a
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users drawing board which has size 1-10,000 in x and 1-10,000 in y.
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Plots for each option are drawn in a window defined by x0,y0 and
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xlength,ylength. Where x0,y0 is the position of the bottom left hand
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corner of the window, and xlength is the width of the window and
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ylength the height of the window.
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--------------------------------------------------------- 10,000
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1 1
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1 -------------------------------------- ^ 1
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1 1 1 1 1
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1 1 1 1 1
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1 1 1 ylength 1
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1 1 1 1 1
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1 1 1 1 1
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1 -------------------------------------- v 1
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1 x0,y0^ 1
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1 <---------------xlength--------------> 1
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--------------------------------------------------------- 1
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1 10,000
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All values are in drawing board units (i.e. 1-10,000, 1-10,000).
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The default window positions are read from a file "ANALMARG" when
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the program is started. Users can have their own file if required.
|
|
As all the plots start at the same position in x and have the same
|
|
width, x0 and xlength are the same for all options. Generally users
|
|
will only want to change the start level of the window y0 and its
|
|
height ylength. This option allows users to change window positions
|
|
whilst running the program. The routine prompts first for the
|
|
number of the option that the users wishes to reposition; then for
|
|
the y start and height; then for the x start and length. Note that
|
|
changes to the x values affect all options. If the user types only
|
|
carriage return for any value it will remain unchanged. The cross-
|
|
hair can be used to choose suitable heights.
|
|
@15. TX 2 @Label a diagram
|
|
|
|
This routine allows users to label any diagrams they have
|
|
produced. They are asked to type in a label. When the user types
|
|
carriage return to finish typing the label the cross-hair appears on
|
|
the screen. The user can position it anywhere on the screen. If the
|
|
user types R (for right justify) the label will be written on the
|
|
diagram with its right end at the cross-hair position. If the user
|
|
types L (for left justify) the label will be written on the diagram
|
|
with its left end at the cross hair position. The cross-hair will
|
|
then immediately reappear. The user may put the same label on
|
|
another part of the diagram as before or if he hits the space bar he
|
|
will be asked if he wishes to type in another label.
|
|
@16. TX 2 @Display a map
|
|
|
|
It is often convenient to plot a map alongside graphed
|
|
analysis in order to indicate features within the sequence. This
|
|
function allows users to draw maps using files arranged in the form
|
|
of EMBL feature tables. Of course the EMBL table are usually only
|
|
used for nucleic acid sequence annotation but, as long as the
|
|
features are written in the correct format, they can be employed by
|
|
this routine. The map is composed of a line representing the
|
|
sequence and then further lines denoting the endpoints of each
|
|
feature the user identifies. The user is asked to define height at
|
|
which the line representing the sequence should be drawn; then for
|
|
the feature height; then for the features to plot.
|
|
@17. TX 1 5 @Short sequence search
|
|
|
|
This routine is used to search for exact matches to short
|
|
sequences. It is equivalent to the restriction enzyme search in
|
|
program NIP. It and can either list matches or present the results
|
|
graphically.
|
|
|
|
Select from searching, screen clearing or file listing. Choose
|
|
a file of strings and the mode of output required.
|
|
|
|
The files of short sequences (strings) and their names need to
|
|
be arranged in a particular way. For example
|
|
ACID/D/E//
|
|
BASIC/R/K/H//
|
|
HYDRO/F/L/I/V/Y//
|
|
GLYCO/N-S/N-T//
|
|
+/R/K/H//
|
|
-/D/E//
|
|
defines various groups of amino acids. Each string or set of
|
|
strings must be preceded by a name, each string must be preceded and
|
|
terminated with a slash (/), and each set of strings by 2 slashes.
|
|
These collections of strings and their names can be read from disk
|
|
or entered from the keyboard. Two files containing sequences are
|
|
currently available. One contains named groups of amino acids. The
|
|
other simply contains the names of all amino acids and gives a
|
|
convenient way of producing a plot of the positions of all the
|
|
different amino acids in the sequence. The user can select strings
|
|
by name from these collections. Results can be displayed name by
|
|
name or all together. Strings entered from the keyboard need to be
|
|
separated by slash characters(/). For the name by name search the
|
|
output looks like:
|
|
MATCHES= 12
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
ACID E 7 7 1
|
|
ACID E 10 3 1
|
|
ACID E 24 14 1
|
|
ACID E 28 4 1
|
|
ACID D 36 8 1
|
|
ACID D 46 10 2
|
|
ACID E 51 5 2
|
|
ACID E 67 16 2
|
|
ACID D 69 2 2
|
|
ACID D 81 12 2
|
|
ACID E 84 3 2
|
|
ACID E 96 12 3
|
|
MATCHES= 10
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
BASIC K 13 13 1
|
|
BASIC R 15 2 1
|
|
BASIC H 26 11 1
|
|
BASIC R 40 14 1
|
|
BASIC H 42 2 2
|
|
BASIC R 59 17 2
|
|
BASIC R 68 9 2
|
|
BASIC K 87 19 2
|
|
BASIC K 89 2 2
|
|
BASIC R 93 4 2
|
|
MATCHES= 1
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
GLYCO NST 4 4 3
|
|
|
|
or when the results are ordered only on position the output looks like:
|
|
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
GLYCO NST 4 3
|
|
ACID E 7 3
|
|
ACID E 10 3
|
|
BASIC K 13 3
|
|
BASIC R 15 2
|
|
ACID E 24 9
|
|
BASIC H 26 2
|
|
ACID E 28 2
|
|
ACID D 36 8
|
|
BASIC R 40 4
|
|
BASIC H 42 2
|
|
ACID D 46 4
|
|
ACID E 51 5
|
|
BASIC R 59 8
|
|
Graphical output marks the position of each string by a short
|
|
vertical line and gives its name at the left end of the line. If the
|
|
top of the screen is reached the program gives the user the
|
|
oportunity to take a hard copy and then will clear the screen and
|
|
restart plotting results at the original start position. Note that
|
|
any character in the string that is not a recognisable protein
|
|
symbol will be treated as a wild card character will match with all
|
|
characters in the searched sequence.
|
|
|
|
Typical dialogue follows.
|
|
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=17
|
|
Search for short sequences
|
|
X 1 Search
|
|
2 List enzyme file
|
|
3 Clear text
|
|
4 Clear graphics
|
|
? 0,1,2,3,4 =2
|
|
1 All acids
|
|
X 2 Named groups
|
|
3 Personal file
|
|
4 Keyboard
|
|
? 0,1,2,3,4 =
|
|
|
|
ACID/D/E//
|
|
BASIC/R/K/H//
|
|
HYDRO/F/L/I/V/Y//
|
|
GLYCO/N-S/N-T//
|
|
+/R/K/H//
|
|
-/D/E//
|
|
DIBASIC/RR/KK/RK/KR//
|
|
TURN/N/D/G/P/S//
|
|
BLOCK/A/Q/E/I/L/M/F/W/V//
|
|
INDIF/R/C/H/K/T/Y//
|
|
End of file
|
|
|
|
|
|
X 1 Search
|
|
2 List enzyme file
|
|
3 Clear text
|
|
4 Clear graphics
|
|
? 0,1,2,3,4 =
|
|
|
|
1 All acids
|
|
X 2 Named groups
|
|
3 Personal file
|
|
4 Keyboard
|
|
? 0,1,2,3,4 =
|
|
|
|
? (y/n) (y) All names n
|
|
? Name=acid
|
|
? Name=basic
|
|
? Name=glyco
|
|
? Name=
|
|
|
|
? (y/n) (y) Show results name by name
|
|
? (y/n) (y) List matches
|
|
|
|
searching
|
|
matches= 59
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
ACID E 7 7 1
|
|
ACID E 10 3 1
|
|
ACID E 24 14 1
|
|
ACID E 28 4 1
|
|
ACID D 36 8 1
|
|
ACID D 46 10 2
|
|
ACID E 51 5 2
|
|
ACID E 67 16 2
|
|
ACID D 69 2 2
|
|
ACID D 81 12 2
|
|
ACID E 84 3 2
|
|
ACID E 96 12 3
|
|
ACID D 116 20 3
|
|
matches= 61
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
BASIC K 13 13 1
|
|
BASIC R 15 2 1
|
|
BASIC H 26 11 1
|
|
BASIC R 40 14 1
|
|
BASIC H 42 2 2
|
|
BASIC R 59 17 2
|
|
...etc
|
|
matches= 2
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
GLYCO NST 4 4 3
|
|
GLYCO NQT 487 483 28
|
|
28 483
|
|
|
|
|
|
X 1 Search
|
|
2 List enzyme file
|
|
3 Clear text
|
|
4 Clear graphics
|
|
? 0,1,2,3,4 =
|
|
|
|
1 All acids
|
|
X 2 Named groups
|
|
3 Personal file
|
|
4 Keyboard
|
|
? 0,1,2,3,4 =
|
|
|
|
? (y/n) (y) Selected names
|
|
|
|
? Name=basic
|
|
? Name=glyco
|
|
? Name=
|
|
|
|
? (y/n) (y) Show results name by name n
|
|
? (y/n) (y) List matches
|
|
|
|
searching
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
GLYCO NST 4 3
|
|
BASIC K 13 9
|
|
BASIC R 15 2
|
|
BASIC H 26 11
|
|
BASIC R 40 14
|
|
BASIC H 42 2
|
|
BASIC R 59 17
|
|
BASIC R 68 9
|
|
BASIC K 87 19
|
|
...etc
|
|
BASIC R 477 14
|
|
BASIC H 479 2
|
|
GLYCO NQT 487 8
|
|
BASIC K 499 12
|
|
BASIC K 501 2
|
|
BASIC K 508 7
|
|
7
|
|
|
|
X 1 Search
|
|
2 List enzyme file
|
|
3 Clear text
|
|
4 Clear graphics
|
|
? 0,1,2,3,4 =
|
|
1 All acids
|
|
X 2 Named groups
|
|
3 Personal file
|
|
4 Keyboard
|
|
? 0,1,2,3,4 =4
|
|
Define search strings by typing a string name
|
|
followed by the string(s)
|
|
? Name=MARY
|
|
? String(s)=AL/VI
|
|
? Name=
|
|
? (y/n) (y) All names
|
|
? (y/n) (y) Show results name by name
|
|
? (y/n) (y) List matches
|
|
|
|
searching
|
|
matches= 12
|
|
NAME SEQUENCE POSITION FRAGMENT LENGTHS
|
|
MARY VI 38 38 10
|
|
MARY AL 63 25 13
|
|
MARY VI 136 73 16
|
|
MARY AL 177 41 19
|
|
MARY AL 217 40 25
|
|
MARY AL 233 16 37
|
|
MARY AL 243 10 40
|
|
MARY AL 256 13 41
|
|
MARY AL 326 70 45
|
|
MARY VI 345 19 51
|
|
MARY AL 396 51 70
|
|
MARY AL 470 74 73
|
|
|
|
|
|
@18. TX 1 5 @Compare a sequence
|
|
|
|
This routine slides a short sequence along the current
|
|
sequence and finds all positions at which a given percentage of the
|
|
amino acids match. Output is in both graphical and listed forms.
|
|
|
|
If users call for dialogue when the routine is selected they
|
|
will be given the choice of keyboard or file input. Define the
|
|
string, and the percentage match. Matches will be plotted out and
|
|
then the user can select to have them listed. Then the routine
|
|
cycles around.
|
|
|
|
The routine slides the search string along the sequence and
|
|
marks the positions at which a minimum percentage score is reached.
|
|
The graphical output draws a vertical line at the match position;
|
|
the height of the line represents the percentage score, so that if
|
|
the line reaches the top of the box the score is 100%.
|
|
|
|
Typical dialogue follows.
|
|
|
|
? Menu or option number=18
|
|
Find percentage matches
|
|
? (y/n) (y) Keep picture
|
|
|
|
? String=aaa
|
|
? Percent match (1.00-100.00) (70.00) =
|
|
|
|
missing graphics
|
|
|
|
Total scoring positions above 70.000 percent = 19
|
|
Scores 2 2 2 2 2 2 2 2 2 2
|
|
Positions 61 131 177 217 226 231 232 267 300 301
|
|
|
|
? Number to list (0-19) (0) =3
|
|
|
|
61
|
|
AIA
|
|
* *
|
|
aaa
|
|
1
|
|
|
|
131
|
|
AIA
|
|
* *
|
|
aaa
|
|
1
|
|
|
|
177
|
|
ALA
|
|
* *
|
|
aaa
|
|
1
|
|
? (y/n) (y) Keep picture n
|
|
|
|
Default String=aaa
|
|
? String=!
|
|
|
|
@19. TX 1 5 @Compare a sequence using a score matrix
|
|
|
|
This routine slides a short sequence along the current
|
|
sequence and finds all positions at which a given level of
|
|
similarity (a cutoff score) is reached. The score is defined by use
|
|
of a score matrix (MDM78). Output is in both graphical and listed
|
|
forms.
|
|
|
|
If users call for dialogue when the routine is selected they
|
|
will be given the choice of keyboard or file input. Define the
|
|
string and the cutoff score. Matches will be plotted out and then
|
|
the user can select to have them listed. Then the routine cycles
|
|
around.
|
|
|
|
The routine slides the search string along the sequence and
|
|
marks the positions at which a the cutoff score is achieved. The
|
|
graphical output draws a vertical line at the match position; the
|
|
height of the line represents the score, so that if the line
|
|
reaches the top of the box the score is the maximum possible.
|
|
|
|
Typical dialogue follows.
|
|
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=19
|
|
Find matches using a score matrix
|
|
? (y/n) (y) Keep picture
|
|
|
|
? String=aaa
|
|
Minimum score= 12 Maximum score= 36
|
|
? Score (12-36) (36) =
|
|
|
|
missing graphics
|
|
|
|
For score 24 the number of matches= 507
|
|
scores 35 35 35 34 34 34 34 34 34 34
|
|
positions 226 231 379 112 133 202 227 267 378
|
|
380
|
|
|
|
? Number to list (0-507) (0) =3
|
|
|
|
226
|
|
ATA
|
|
* *
|
|
aaa
|
|
1
|
|
|
|
231
|
|
SAA
|
|
**
|
|
aaa
|
|
1
|
|
|
|
379
|
|
GAA
|
|
**
|
|
aaa
|
|
1
|
|
? (y/n) (y) Keep picture n
|
|
|
|
Default String=aaa
|
|
? String=!
|
|
@20. TX 5 @Search for a motif using a weight matrix
|
|
|
|
This function performs searches for short sequence motifs
|
|
using an appropriate weight matrix. In addition it can be used to
|
|
create or modify weight matrices. In order to perform a search the
|
|
only input required is the name of the file containing the weight
|
|
matrix. The results can be presented graphically or listed. The
|
|
graphical presentation will draw line at the position of any matches
|
|
found; the height of the line is proportional to the score.
|
|
|
|
For a search, select "use weight matrix", supply the name of
|
|
the file containing the weight matrix, and choose between having
|
|
results plotted or listed. If dialogue is requested when the
|
|
function is selected users can alter the cutoff score employed.
|
|
|
|
To create a weight matrix several steps are involved. A file
|
|
containing an alignment of known motifs is required. (This file must
|
|
be created before the current option is selected. The format is a
|
|
follows: each sequence is written on a separate line with at least
|
|
one space at the beginning; each sequence is terminated by a space
|
|
character, and can be followed by a name. The sequences must be
|
|
aligned.) Supply the name of the file of aligned sequences. The
|
|
program reads and displays the sequences. Choose between "summing
|
|
logs of weights" or summing weights (i.e. whether to multiply or add
|
|
weights). If logs are used all scores will be negative. Choose if
|
|
all positions in the set of aligned sequences should be used or if a
|
|
mask should be applied. If so selected, define a mask as a string of
|
|
symbols, in which symbol - means ignore and any other symbol means
|
|
use. E.g. xx-x--abc means use all positions except 3,5 and 6.
|
|
|
|
The program will calculate weights as the frequencies of each
|
|
amino acid at each unmasked position in the set of aligned
|
|
sequences. These weights are then applied to the set of aligned
|
|
sequences to give a range of "observed" scores. The mean and
|
|
standard deviation of these scores is displayed. The user is asked
|
|
to supply several values to be used when the weight matrix is
|
|
applied to other sequences: a cutoff score (by default, the mean
|
|
minus 3 standard deviations), a top score for scaling graphical
|
|
results (by default, the mean plus 3 standard deviations), and a
|
|
position to identify (this means that if a particular amino acid
|
|
within the motif is used as a "landmark", such as the G of the
|
|
helix-turn-helix motif, then its position will be marked in plots).
|
|
All these values are stored along with the weight matrix. Finally
|
|
supply the name of a file to contain the weight matrix.
|
|
|
|
Weight matrices can be "rescaled" using a set of aligned
|
|
sequences in much the same ways as a matrix is created. The purpose
|
|
is to redefine the cutoff scores, and rescaling does not alter any
|
|
other values in the weight matrix file.
|
|
|
|
The methods have changed considerably but were first outlined
|
|
in Staden, R. Nucl. Acid Res. 12 505-519 1984, and Staden, R.
|
|
Genetic engineering: principles and methods vol 7, Edited by J.K.
|
|
Setlow and A. Hollaender, Plenum publishing corp., 1985.
|
|
|
|
The methods have always had to deal with the problem of zeroes
|
|
in the matrices. The current versions employ "Laplaces Law of
|
|
Succession" in which 1 is added to each term.
|
|
|
|
It is now possible to apply a mask to a set of aligned
|
|
sequences in order to give weight to selected positions only.
|
|
Sequences have superimposed functions: some parts may be of general
|
|
structural importance and give rise to an overall framework, and
|
|
other parts give specificity and hence are not common; we may want
|
|
to use a set of aligned sequences to define a motif, but want to use
|
|
only the framework positions. Alternatively we may want to pick out
|
|
only those parts of a set of aligned sequences that give a
|
|
particular property, and to ignore other similarities that are due
|
|
to some other property and which could obscure the pattern we are
|
|
interested in. The ability to define a mask allows certain positions
|
|
to be used in the motif and others to be ignored, and yet still
|
|
permits the use of a set of aligned sequences to calculate weights.
|
|
|
|
Typical dialogue is shown below.
|
|
? Menu or option number=20
|
|
X 1 Use weight matrix
|
|
2 Make weight matrix
|
|
3 Rescale weight matrix
|
|
? 0,1,2,3 =2
|
|
? Name of aligned sequences file=[rs.motifs]hth.seq
|
|
1 QESVADKMGMGQSGVGALFN LAMBDA.REP
|
|
2 QTKTAKDLGVYQSAINKAIH LAMBDA.CRO
|
|
3 QAALGKMVGVSNVAISQWQR P22.REP
|
|
4 QRAVAKALGISDAAVSQWKE P22.CRO
|
|
5 QAELAQKVGTTQQSIEQLEN 434.REP
|
|
6 QTELATKAGVKQQSIQLIEA 434.CRO
|
|
7 RQEIGQIVGCSRETVGRILK CAP
|
|
8 RGDIGNYLGLTVETISRLLG Fnr
|
|
9 LYDVAEYAGVSYQTVSRVVN LAC.R
|
|
10 IKDVARLAGVSVATVSRVIN GAL.R
|
|
11 TEKTAEAVGVDKSQISRWKR LAMBDA.CII
|
|
12 QRKVADALGINESQISRWKG P22.CI
|
|
13 KEEVAKKCGITPLQVRVWCN MAT.ALPHA
|
|
14 TRKLAQKLGVEQPTLYWHVK TETR.TN10
|
|
15 TRRLAERLGVQQPALYWHFK TETR.pSC1
|
|
16 QRELKNELGAGIATITRGSN TRP.REP
|
|
17 RQQLAIIFGIGVSTLYRYFP H-INVERSN
|
|
18 ATEIAHQLSIARSTVYKILE TN3.RESOL
|
|
19 ASHISKTMNIARSTVYKVIN GD.RESOLV
|
|
20 IASVAQHVCLSPSRLSHLFR ARA.C
|
|
21 RAEIAQRLGFRSPNAAEEHL LEX.R
|
|
Length of motif 20
|
|
? (y/n) (y) Sum logs of weights
|
|
? (y/n) (y) Use all motif positions n
|
|
x means use, - means ignore
|
|
e.g. xx-x---x-x means use positions 1,2,4,8,10
|
|
? Mask=--xxxxxxxxxxxx------
|
|
Applying weights to input sequences
|
|
1 -57.143 QESVADKMGMGQSGVGALFN
|
|
2 -55.087 QTKTAKDLGVYQSAINKAIH
|
|
3 -58.079 QAALGKMVGVSNVAISQWQR
|
|
4 -54.986 QRAVAKALGISDAAVSQWKE
|
|
5 -55.181 QAELAQKVGTTQQSIEQLEN
|
|
6 -55.874 QTELATKAGVKQQSIQLIEA
|
|
7 -56.692 RQEIGQIVGCSRETVGRILK
|
|
8 -57.722 RGDIGNYLGLTVETISRLLG
|
|
9 -55.363 LYDVAEYAGVSYQTVSRVVN
|
|
10 -55.769 IKDVARLAGVSVATVSRVIN
|
|
11 -56.786 TEKTAEAVGVDKSQISRWKR
|
|
12 -55.833 QRKVADALGINESQISRWKG
|
|
13 -56.279 KEEVAKKCGITPLQVRVWCN
|
|
14 -53.125 TRKLAQKLGVEQPTLYWHVK
|
|
15 -55.833 TRRLAERLGVQQPALYWHFK
|
|
16 -58.651 QRELKNELGAGIATITRGSN
|
|
17 -56.749 RQQLAIIFGIGVSTLYRYFP
|
|
18 -56.986 ATEIAHQLSIARSTVYKILE
|
|
19 -60.618 ASHISKTMNIARSTVYKVIN
|
|
20 -58.988 IASVAQHVCLSPSRLSHLFR
|
|
21 -58.002 RAEIAQRLGFRSPNAAEEHL
|
|
Top score -53.125 Bottom score -60.618
|
|
Mean -56.655 Standard deviation 1.617
|
|
Mean minus 3.sd -61.505 Mean plus 3.sd -51.804
|
|
? Cutoff score (-999.00-9999.00) (-61.51) =
|
|
? Top score for scaling plots (-61.51-999.00) (-51.80) =
|
|
? Position to identify (0-20) (1) =9
|
|
? Title=hth
|
|
? Name for new weight matrix file=1.wts
|
|
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=20
|
|
X 1 Use weight matrix
|
|
2 Make weight matrix
|
|
3 Rescale weight matrix
|
|
? 0,1,2,3 =
|
|
|
|
? Motif weight matrix file=1.wts
|
|
hth
|
|
? (y/n) (y) Use frequencies as weights
|
|
? (y/n) (y) Plot results n
|
|
5 -61.46 STEISELIKQRIAQFNVVSE
|
|
13 -58.93 KQRIAQFNVVSEAHNEGTIV
|
|
21 -60.42 VVSEAHNEGTIVSVSDGVIR
|
|
57 -59.39 GNRYAIALNLERDSVGAVVM
|
|
59 -61.47 RYAIALNLERDSVGAVVMGP
|
|
79 -59.90 YADLAEGMKVKCTGRILEVP
|
|
88 -61.41 VKCTGRILEVPVGRGLLGRV
|
|
104 -60.38 LGRVVNTLGAPIDGKGPLDH
|
|
127 -60.13 SAVEAIAPGVIERQSVDQPV
|
|
129 -59.91 VEAIAPGVIERQSVDQPVQT
|
|
133 -60.79 APGVIERQSVDQPVQTGYKA
|
|
139 -61.12 RQSVDQPVQTGYKAVDSMIP
|
|
175 -58.90 KTALAIDAIINQRDSGIKCI
|
|
191 -60.95 IKCIYVAIGQKASTISNVVR
|
|
195 -60.94 YVAIGQKASTISNVVRKLEE
|
|
215 -60.66 HGALANTIVVVATASESAAL
|
|
254 -60.56 EDALIIYDDLSKQAVAYRQI
|
|
260 -60.08 YDDLSKQAVAYRQISLLLRR
|
|
297 -61.00 LLERAARVNAEYVEAFTKGE
|
|
314 -61.29 KGEVKGKTGSLTALPIIETQ
|
|
330 -60.49 IETQAGDVSAFVPTNVISIT
|
|
363 -57.63 GIRPAVNPGISVSRVGGAAQ
|
|
365 -61.48 RPAVNPGISVSRVGGAAQTK
|
|
371 -61.02 GISVSRVGGAAQTKIMKKLS
|
|
382 -57.90 QTKIMKKLSGGIRTALAQYR
|
|
394 -60.07 RTALAQYRELAAFSQFASDL
|
|
424 -59.95 GQKVTELLKQKQYAPMSVAQ
|
|
430 -58.89 LLKQKQYAPMSVAQQSLVLF
|
|
432 -61.14 KQKQYAPMSVAQQSLVLFAA
|
|
438 -58.58 PMSVAQQSLVLFAAERGYLA
|
|
458 -61.06 DVELSKIGSFEAALLAYVDR
|
|
466 -61.00 SFEAALLAYVDRDHAPLMQE
|
|
483 -60.48 MQEINQTGGYNDEIEGKLKG
|
|
494 -60.61 DEIEGKLKGILDSFKATQSW
|
|
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=d20
|
|
X 1 Use weight matrix
|
|
2 Make weight matrix
|
|
3 Rescale weight matrix
|
|
? 0,1,2,3 =
|
|
|
|
? Motif weight matrix file=1.wts
|
|
hth
|
|
? (y/n) (y) Use frequencies as weights
|
|
? Cutoff score (-9999.00-9999.00) (-61.51) =-56.
|
|
? (y/n) (y) Plot results n
|
|
|
|
|
|
@21. TX 3 @Calculate amino acid composition
|
|
|
|
This function calculates the amino acid composition and
|
|
molecular weight for the active region.
|
|
? Menu or option number=21
|
|
Sequence composition
|
|
|
|
A C S T P A G N D E Q B Z H
|
|
N 3. 32. 23. 18. 57. 47. 16. 28. 31. 28. 0. 0. 7.
|
|
% 0.6 6.2 4.5 3.5 11.1 9.1 3.1 5.4 6.0 5.4 0.0 0.0 1.4
|
|
W 309. 2786. 2325. 1748. 4051. 2682. 1826. 3222. 4003. 3588. 0. 0.
|
|
960.
|
|
|
|
A R K M I L V F Y W - X ?
|
|
N 30. 24. 11. 40. 47. 41. 14. 15. 1. 0. 0. 0. 1.
|
|
% 5.8 4.7 2.1 7.8 9.1 8.0 2.7 2.9 0.2 0.0 0.0 0.0 0.2
|
|
W 4686. 3076. 1443. 4527. 5319. 4065. 2060. 2448. 186. 0. 0. 0.
|
|
0.
|
|
Total molecular weight= 55328.
|
|
|
|
@22. TX 3 4 @Plot hydrophobicity
|
|
|
|
This routine plots the hydrophobicity of each section of the
|
|
sequence using the hydrophobicity values of Kyte and Doolittle (J.
|
|
Mol. Biol. 157, 105-132 (1982)). A window of size span is slid
|
|
along the sequence and a sum calculated for each position.
|
|
|
|
If dialogue is requested select a span length and a plot
|
|
interval.
|
|
|
|
The diagrams are on the same scale as Fig. 6 of the Kyte and
|
|
Doolittle paper and values of + and - 50 could be assigned to the
|
|
top and bottom of the diagram with corresponding values in between
|
|
(-40,-20,0,20,40 are shown in the paper).
|
|
? Menu or option number=d22
|
|
Plot hydrophobicity
|
|
? odd span length (1-101) (11) =
|
|
? plot interval (1-101) (3) =
|
|
|
|
missing graphics
|
|
@23. TX 3 4 @Plot charge
|
|
|
|
This routine plots the charge of each section of the sequence.
|
|
A window of size span is slid along the sequence and a sum
|
|
calculated for each position. Amino acids are assigned charges of 1,
|
|
-1 or 0.
|
|
|
|
If dialogue is requested select a span length and a plot
|
|
interval.
|
|
|
|
Typical dialogue follows.
|
|
|
|
? Menu or option number=d23
|
|
Plot charge
|
|
? odd span length (1-101) (11) =
|
|
? plot interval (1-101) (3) =
|
|
|
|
missing graphics
|
|
|
|
@24. TX 4 @Plot robson prediction
|
|
|
|
This routine uses the method of Garnier J, Osguthorpe D J, and
|
|
Robson B. (1978) J. Mol. Biol. 120, 97-120 to predict secondary
|
|
structures. The method divides protein secondary structures into 4
|
|
classes: helix, extended (usually referred to as sheet), turn and
|
|
coil. The routine calculates the likelihood that each segment of the
|
|
sequence lies in each of these classes. Results are presented
|
|
graphically or listed.
|
|
|
|
If dialogue is requested choose between plotted or listed
|
|
output.
|
|
|
|
Each residue has a certain probability of being found in each
|
|
of the 4 classes. This probability depends both on its own amino
|
|
acid type and also the 8 amino acids found to either side along the
|
|
protein chain. Four tables of weights, each 20 by 17 elements are
|
|
used to calculate the likelihood that each residue along the chain
|
|
falls into one of the four classes of structure. The most likely
|
|
structure at each point is the one with the highest score. The four
|
|
values are plotted in strips labelled H, E, T and C. Below, a strip
|
|
labelled D for decision is divided into four levels, each
|
|
corresponding to one of the four structure types. Their top to
|
|
bottom order is the same as that for the strips above, i.e C, T, E,
|
|
and H. For each residue the program measures which of the four
|
|
likelhoods is highest. It places a single dot at the mid-point of
|
|
the corresponding strip, and also at the appropriate level in the
|
|
strip labelled D.
|
|
|
|
It should be noted that the method, when tested by Kabsch W
|
|
and Sander C, (1983) Febs. Lett. 155 (179-182), although one of the
|
|
better ones, was correct for only about 56% of residues.
|
|
|
|
Typical dialogue follows.
|
|
? Menu or option number=d24
|
|
Plot Robson secondary structure predictions
|
|
? (y/n) (y) Plot results n
|
|
|
|
9 S 217 -7 -39 15
|
|
10 E 226 5 -27 -39
|
|
11 L 233 -7 -26 -15
|
|
12 I 229 -23 9 4
|
|
13 K 214 -8 10 -8
|
|
14 Q 178 42 19 5
|
|
15 R 131 54 16 3
|
|
16 I 86 42 -31 -23
|
|
17 A 55 52 -30 -15
|
|
18 Q 15 67 4 25
|
|
19 F -34 86 47 74
|
|
20 N -41 74 17 106
|
|
21 V -16 118 -5 100
|
|
22 V 64 88 5 115
|
|
23 S 96 38 26 155
|
|
24 E 133 -25 13 96
|
|
25 A 118 -98 25 100
|
|
26 H 110 -150 37 86
|
|
27 N 57 -201 37 66
|
|
28 E 51 -140 11 -4
|
|
29 G 2 -77 37 9
|
|
30 T 2 28 28 7
|
|
31 I -11 117 -21 22
|
|
32 V -23 178 -55 5
|
|
33 S -54 193 -14 35
|
|
34 V -46 123 5 30
|
|
35 S -54 53 51 80
|
|
36 D -60 1 86 55
|
|
37 G -66 8 57 49
|
|
38 V -1 128 -30 -5
|
|
39 I 11 212 -56 -33
|
|
40 R 16 204 -44 -57
|
|
...etc
|
|
|
|
@26. TX 4 @Draw a helix wheel
|
|
|
|
A helical representation of segments of the sequence is shown.
|
|
The display includes a schematic of the helix showing the links
|
|
between residues, with each vertex numbered according to position;
|
|
the sequence element at each vertex; a symbol denoting a
|
|
classification as hydrophobic(.), positively charged(+), negatively
|
|
charged(-), or otherwise( ). The residue number of the first
|
|
sequence element in the current window is displayed at the top-
|
|
left-hand corner of the diagram. Also at the top-left corner the
|
|
sequence in the current window is listed. Below this is the total
|
|
hydrophobicity and hydrophobic moment for the window calculated
|
|
according to Eisenberg et al J. Mol. Biol. 179, 125-142 (1984).
|
|
|
|
If dialogue is requested the user is asked for the angle to
|
|
define the turn between residues as seen looking along the helix,
|
|
and a window length. The window length can be up to 60, with default
|
|
18, and the angle has a default of 100 degrees. Note that 18 x 100
|
|
is 5 turns. When the option is selected the first segment in the
|
|
current active region is displayed then the bell rings. If the user
|
|
types only return, the display will click on by one residue; if
|
|
another number is typed, say N, then the display will click forwards
|
|
(or backwards if N is negative) by N residues. If the wheel runs off
|
|
either end of the sequence the option will be exited.
|
|
|
|
Typical dialogue follows.
|
|
? Menu or option number=d26
|
|
? Angle (1-130) (100) =
|
|
? Window (1-60) (18) =
|
|
|
|
missing graphics
|
|
|
|
@25. TX 3 4 @Plot hydrophobic moment
|
|
|
|
This routine plots hydrophobic moment and hydrophobicity
|
|
according to Eisenberg et al J. Mol. Biol. 179, 125-142 (1984). The
|
|
mean hydrophobicity per residue in the window is plotted on a scale
|
|
-1.0 to 1.5, and the mean hydrophobic moment per residue on a scale
|
|
0.0 to 1.5. The hydrophobicity is shown in the top frame with the
|
|
hydrophobic moment below. The plot is arranged so that the value
|
|
shown at position x represents the mean value for residues x-
|
|
window+1 to x, where window is the window length.
|
|
|
|
If dialogue is requested the user can select a window length,
|
|
and the angle used for the hydrophobic moment calculation.
|
|
|
|
Note that according to Eisenberg et al, in transmembrane
|
|
proteins an "initiator" is required. This is either a very
|
|
hydrophobic single helix with <H> >=0.68, or a moderately
|
|
hydrophobic pair of helices whose <H> sum to >= 1.1. Other helices
|
|
are then accepted as transmembrane if their <H> >= 0.42
|
|
|
|
The following rules are claimed: if <H> < 0.51 and points lie
|
|
below the line <M> = -0.392 + 0.603x <H> they are "globular", if
|
|
they lie above this line they are "surface". If <H> > 0.51 and they
|
|
lie above the line <M> = 0.6 - 0.342x<H> they are "monomeric", if
|
|
above "multimeric".
|
|
|
|
Typical dialogue follows.
|
|
|
|
? Menu or option number=d25
|
|
? Angle (1-130) (100) =
|
|
? Window (1-60) (18) =
|
|
? Plot interval (1-101) (3) =
|
|
|
|
missing graphics
|
|
|
|
|
|
@27. TX 1 @Back translate to dna
|
|
|
|
This routine back translates protein sequences into DNA using
|
|
the standard genetic code. The level of redundancy can be plotted
|
|
and the backtranslation saved to a file.
|
|
|
|
The translation can use either the IUB symbols shown below, or
|
|
a set of codon preferences. If a set of codon preferences are used
|
|
they must conform to the format of codon tables produced by the
|
|
nucleotide analysis program, and the back translation will contain
|
|
the favoured codons. If there is no favoured codon the IUB symbols
|
|
will be employed. The window length for plotting the redundancy is
|
|
in codons.
|
|
|
|
The program will plot the redundancy along the sequence and
|
|
hence can be used to find the best sequences to use as primers. Note
|
|
that the program plots the inverse, and so the higher the plot the
|
|
LESS redundant the sequence. For primers look for peaks rather than
|
|
troughs.
|
|
|
|
The DNA sequence can be saved to a file and analysed using the
|
|
nucleotide analysis program. Depending on the application it is
|
|
often useful to produce a back translation using both a table of
|
|
codon preferences and one using the IUB symbols. This is because the
|
|
restriction enzyme search program can distinguish between definite
|
|
and possible cuts in the sequence. These matches are what the
|
|
program terms "definite matches" and are ones in which the
|
|
specification of the recognition sequence corresponds exactly to
|
|
that of the back translation. The program will also find what it
|
|
terms "possible matches" which are ones that depend on the
|
|
particular codons chosen for each amino acid. These are sites at
|
|
which recognition sequences could be engineered to produce a cut in
|
|
the DNA without changing the amino acid, but which are not
|
|
necessarily found in the original sequence.
|
|
|
|
|
|
NC-IUB SYMBOLS
|
|
|
|
A,C,G,T
|
|
R (A,R) 'puRine'
|
|
Y (T,C) 'pYrimidine'
|
|
W (A,T) 'Weak'
|
|
S (C,G) 'Strong'
|
|
M (A,C) 'aMino'
|
|
K (G,T) 'Keto'
|
|
H (A,T,C) 'not G'
|
|
B (G,C,T) 'not A'
|
|
V (G,A,C) 'not T'
|
|
D (G,A,T) 'not C'
|
|
N (G,A,C,T) 'aNy'
|
|
|
|
Typical dialogue follows.
|
|
|
|
? Menu or option number=d27
|
|
Back translate
|
|
? (y/n) (y) No codon preference
|
|
? (y/n) (y) Plot redundancy n
|
|
? (y/n) (y) Save DNA to disk
|
|
? File name for DNA sequence=tt:
|
|
ATGCARYTNAAYWSNACNGARATHWSNGARYTNATHAARCARMGNATHGCNCARTTYAAY
|
|
GTNGTNWSNGARGCNCAYAAYGARGGNACNATHGTNWSNGTNWSNGAYGGNGTNATHMGN
|
|
ATHCAYGGNYTNGCNGAYTGYATGCARGGNGARATGATHWSNYTNCCNGGNAAYMGNTAY
|
|
GCNATHGCNYTNAAYYTNGARMGNGAYWSNGTNGGNGCNGTNGTNATGGGNCCNTAYGCN
|
|
GAYYTNGCNGARGGNATGAARGTNAARTGYACNGGNMGNATHYTNGARGTNCCNGTNGGN
|
|
MGNGGNYTNYTNGGNMGNGTNGTNAAYACNYTNGGNGCNCCNATHGAYGGNAARGGNCCN
|
|
YTNGAYCAYGAYGGNTTYWSNGCNGTNGARGCNATHGCNCCNGGNGTNATHGARMGNCAR
|
|
WSNGTNGAYCARCCNGTNCARACNGGNTAYAARGCNGTNGAYWSNATGATHCCNATHGGN
|
|
MGNGGNCARMGNGARYTNATHATHGGNGAYMGNCARACNGGNAARACNGCNYTNGCNATH
|
|
GAYGCNATHATHAAYCARMGNGAYWSNGGNATHAARTGYATHTAYGTNGCNATHGGNCAR
|
|
AARGCNWSNACNATHWSNAAYGTNGTNMGNAARYTNGARGARCAYGGNGCNYTNGCNAAY
|
|
ACNATHGTNGTNGTNGCNACNGCNWSNGARWSNGCNGCNYTNCARTAYYTNGCNMGNATG
|
|
CCNGTNGCNYTNATGGGNGARTAYTTYMGNGAYMGNGGNGARGAYGCNYTNATHATHTAY
|
|
GAYGAYYTNWSNAARCARGCNGTNGCNTAYMGNCARATHWSNYTNYTNYTNMGNMGNCCN
|
|
CCNGGNMGNGARGCNTTYCCNGGNGAYGTNTTYTAYYTNCAYWSNMGNYTNYTNGARMGN
|
|
GCNGCNMGNGTNAAYGCNGARTAYGTNGARGCNTTYACNAARGGNGARGTNAARGGNAAR
|
|
ACNGGNWSNYTNACNGCNYTNCCNATHATHGARACNCARGCNGGNGAYGTNWSNGCNTTY
|
|
GTNCCNACNAAYGTNATHWSNATHACNGAYGGNCARATHTTYYTNGARACNAAYYTNTTY
|
|
AAYGCNGGNATHMGNCCNGCNGTNAAYCCNGGNATHWSNGTNWSNMGNGTNGGNGGNGCN
|
|
GCNCARACNAARATHATGAARAARYTNWSNGGNGGNATHMGNACNGCNYTNGCNCARTAY
|
|
MGNGARYTNGCNGCNTTYWSNCARTTYGCNWSNGAYYTNGAYGAYGCNACNMGNAARCAR
|
|
YTNGAYCAYGGNCARAARGTNACNGARYTNYTNAARCARAARCARTAYGCNCCNATGWSN
|
|
GTNGCNCARCARWSNYTNGTNYTNTTYGCNGCNGARMGNGGNTAYYTNGCNGAYGTNGAR
|
|
YTNWSNAARATHGGNWSNTTYGARGCNGCNYTNYTNGCNTAYGTNGAYMGNGAYCAYGCN
|
|
CCNYTNATGCARGARATHAAYCARACNGGNGGNTAYAAYGAYGARATHGARGGNAARYTN
|
|
AARGGNATHYTNGAYWSNTTYAARGCNACNCARWSNTGG---
|
|
|
|
|
|
@28. TX 5 @Search for patterns of motifs
|
|
|
|
This option searches for patterns of motifs. Patterns can be
|
|
defined interactively or read from files. Results can be displayed
|
|
in several ways in both graphical and textual form. Used to create
|
|
pattern files for searching libraries. The option is extremely
|
|
flexible and consequently the following documentation is quite
|
|
lengthy. However the routine is capable of searching for almost any
|
|
known pattern. In addition the flexibility does not necessitate
|
|
difficulty of use, and the userinterface has been simplified
|
|
considerably since the methods were first published.
|
|
|
|
Users should refer to the "typical dialogue" shown below for
|
|
the most helpful information on using the program.
|
|
|
|
There are currently four ways to display the matching
|
|
patterns: 1=each individual motif and its position is listed; 2=all
|
|
the sequence between, and including the two outermost motifs is
|
|
listed; 3=graphical, with a vertical line marking the position of
|
|
the leftmost motif; 4 = EMBL feature table format, where the KEYNAM
|
|
field is the motif name, the FROM and TO fields denote the ends of
|
|
the match, and the DESCRIPTION field is "Program".
|
|
|
|
When it is defined for the first time a pattern must be
|
|
entered interactively at the keyboard, but the pattern description
|
|
can be saved to a file. This file can be used for all subsequent
|
|
searches.
|
|
|
|
When defining a pattern interactively select a motif class and
|
|
the program will request the required inputs.
|
|
|
|
The program gives each motif an identifying name and number.
|
|
For motifs other than the first, a range of allowed positions must
|
|
be defined (Note that sets of motifs included using the OR operator
|
|
will all be given the same range, and so the program will only
|
|
request range values for the first motif in any such set). To
|
|
specify the allowed range for a motif the user must supply the
|
|
following: the identifying number of the motif, relative to which
|
|
the current motifs positions are to be defined (termed the
|
|
"reference motif"); a "relative start position" and a range. The
|
|
relative start position can be negative or positive. A negative
|
|
start position means that although the reference motif is searched
|
|
for first, the current motif can be found to its left. A zero
|
|
relative start position means their left ends are superimposed. The
|
|
default start position is to butt-joint the motif to righthand end
|
|
of the "reference motif". The range is "the number of extra
|
|
positions" that the motif can take.
|
|
|
|
The program will display the probability of finding each
|
|
motif. These values are presented in the following form: .1234E-5
|
|
means 0.1234 times 10 to the power -5.
|
|
|
|
After the pattern has been defined, the program will type a
|
|
description of it on the screen. It will then allow the user to give
|
|
an overall cutoff score and overall probability cutoff.
|
|
|
|
Typical dialogue for all the different motif classes is
|
|
displayed below.
|
|
|
|
? Menu or option number=28
|
|
Pattern searcher
|
|
? (y/n) (y) Read pattern from keyboard
|
|
X 1 Exact match
|
|
2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =
|
|
? Motif name=aa
|
|
? String=aa
|
|
Probability of score 2.0000 = 0.123E-01
|
|
X 1 Exact match
|
|
2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =2
|
|
? Motif name=pmatch
|
|
X 1 And
|
|
2 Or
|
|
3 Not
|
|
? 0,1,2,3 =
|
|
? Number of reference motif (1-1) (1) =
|
|
? Relative start position (-1000-1000) (3) =
|
|
? Number of extra positions (0-1000) (0) =
|
|
? String=qqq
|
|
? Minimum matches (1.00-3.00) (3.00) =2
|
|
Probability of score 2.0000 = 0.858E-02
|
|
1 Exact match
|
|
X 2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =3
|
|
? Motif name=sm
|
|
X 1 And
|
|
2 Or
|
|
3 Not
|
|
? 0,1,2,3 =
|
|
? Number of reference motif (1-2) (2) =
|
|
? Relative start position (-1000-1000) (4) =
|
|
? Number of extra positions (0-1000) (0) =
|
|
? String=wqa
|
|
? Minimum score (11.00-53.00) (53.00) =36
|
|
Probability of score 36.0000 = 0.531E-02
|
|
1 Exact match
|
|
2 Percentage match
|
|
X 3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =4
|
|
? Motif name=hth
|
|
X 1 And
|
|
2 Or
|
|
3 Not
|
|
? 0,1,2,3 =
|
|
? Number of reference motif (1-3) (3) =
|
|
? Relative start position (-1000-1000) (4) =
|
|
? Number of extra positions (0-1000) (0) =
|
|
? Weight matrix file name=hth
|
|
HELIX TURN HELIX PABO SAUER WEIGHTS 17-11-87
|
|
Probability of score -51.5860 = 0.230E-04
|
|
1 Exact match
|
|
2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
X 4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =5
|
|
? Motif name=repeat
|
|
X 1 And
|
|
2 Or
|
|
3 Not
|
|
? 0,1,2,3 =
|
|
? Number of reference motif (1-4) (4) =
|
|
? Relative start position (-1000-1000) (21) =
|
|
? Number of extra positions (0-1000) (0) =3
|
|
? Repeat length (1-60) (6) =3
|
|
? Minimum gap (0-60) (0) =
|
|
? Maximum gap (0-60) (0) =2
|
|
? Minimum score (11.00-60.00) (36.00) =
|
|
Probability of score 36.0000 = 0.445E-01
|
|
1 Exact match
|
|
2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
X 5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =6
|
|
? Motif name=mset
|
|
X 1 And
|
|
2 Or
|
|
3 Not
|
|
? 0,1,2,3 =
|
|
? Number of reference motif (1-5) (5) =
|
|
? Relative start position (-1000-1000) (1) =
|
|
? Number of extra positions (0-1000) (0) =
|
|
X 1 Keyboard input
|
|
2 File input
|
|
? 0,1,2 =
|
|
Separate sets with commas
|
|
? String=AVL,AST,,WYRF
|
|
? Minimum matches (1.00-4.00) (4.00) =3
|
|
Probability of score 3.0000 = 0.718E-02
|
|
1 Exact match
|
|
2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
X 6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =7
|
|
? (y/n) (y) Save pattern in a file
|
|
? Pattern definition file=EXAM.PAT
|
|
Motif 6 needs a file name to store set as a weight matrix
|
|
? Weight matrix file name=DEMO.WTS
|
|
Weight matrix needs a title
|
|
? Title=Demonstration class 6 weight matrix
|
|
|
|
Pattern description
|
|
|
|
Motif 1 named aa is of class 1
|
|
Which is an exact match to the string
|
|
aa
|
|
Motif 2 named pmatch is of class 2
|
|
which is a match of score 2. to the string
|
|
qqq
|
|
and the N-terminal residue can take positions 3 to 3
|
|
relative to the N-terminal end of motif 1
|
|
It is anded with the previous motif.
|
|
Motif 3 named sm is of class 3
|
|
which is a match of score 36. to the string
|
|
wqa
|
|
and the N-terminal residue can take positions 4 to 4
|
|
relative to the N-terminal end of motif 2
|
|
It is anded with the previous motif.
|
|
Motif 4 named hth is of class 4
|
|
Which is a match to a weight matrix with score -51.586
|
|
and the N-terminal residue can take positions 4 to 4
|
|
relative to the N-terminal end of motif 3
|
|
It is anded with the previous motif.
|
|
Motif 5 named repeat is of class 5
|
|
Which is a repeat with repeat length 3 and score 36.
|
|
The loop-out can have sizes 0 to 2
|
|
and the N-terminal residue can take positions 21 to 24
|
|
relative to the N-terminal end of motif 4
|
|
It is anded with the previous motif.
|
|
Motif 6 named mset is of class 6
|
|
Which is membership of a set with score 3.000
|
|
It is anded with the previous motif.
|
|
Probability of finding pattern = 0.4109E-14
|
|
Expected number of matches = 0.2539E-10
|
|
? Maximum pattern probability (0.00-1.00) (1.00) =
|
|
? Minimum pattern score (-9999.00-9999.00) (-9999.00) =
|
|
Select display mode
|
|
X 1 Motif by motif
|
|
2 Inclusive
|
|
3 Graphical
|
|
4 EMBL feature table
|
|
? 0,1,2,3,4 =
|
|
Searching
|
|
|
|
Total matches found 0
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=6
|
|
Page through text files
|
|
? Name of file to read=exam.pat
|
|
A1 aa Class
|
|
aa
|
|
@ End of string
|
|
A2 pmatch Class
|
|
1 Relative motif
|
|
3 Relative start position
|
|
0 Number of extra positions
|
|
qqq
|
|
@ End of string
|
|
2.00000 Cutoff
|
|
A3 sm Class
|
|
2 Relative motif
|
|
4 Relative start position
|
|
0 Number of extra positions
|
|
wqa
|
|
@ End of string
|
|
36.00000 Cutoff
|
|
A4 hth Class
|
|
3 Relative motif
|
|
4 Relative start position
|
|
0 Number of extra positions
|
|
hth File name
|
|
A5 repeat Class
|
|
4 Relative motif
|
|
21 Relative start position
|
|
3 Number of extra positions
|
|
3 Length
|
|
0 Minimum loop
|
|
2 Maximum loop
|
|
36.00000 Cutoff
|
|
A6 mset Class
|
|
5 Relative motif
|
|
1 Relative start position
|
|
0 Number of extra positions
|
|
DEMO.WTS File name
|
|
End of file
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=6
|
|
Page through text files
|
|
? Name of file to read=demo.wts
|
|
Demonstration class 6 weight matrix
|
|
4 0 3.000 4.000
|
|
P 1 2 3 4
|
|
N 0 0 0 0
|
|
C 0 0 0 0
|
|
S 0 1 0 0
|
|
T 0 1 0 0
|
|
P 0 0 0 0
|
|
A 1 1 0 0
|
|
G 0 0 0 0
|
|
N 0 0 0 0
|
|
D 0 0 0 0
|
|
E 0 0 0 0
|
|
Q 0 0 0 0
|
|
B 0 0 0 0
|
|
Z 0 0 0 0
|
|
H 0 0 0 0
|
|
R 0 0 0 1
|
|
K 0 0 0 0
|
|
M 0 0 0 0
|
|
I 0 0 0 0
|
|
L 1 0 0 0
|
|
V 1 0 0 0
|
|
F 0 0 0 1
|
|
Y 0 0 0 1
|
|
W 0 0 0 1
|
|
End of file
|
|
Menus and their numbers are
|
|
m0 = This menu
|
|
m1 = General
|
|
m2 = Screen control
|
|
m3 = Statistical analysis of content
|
|
m4 = Structure
|
|
m5 = Search
|
|
? = Help
|
|
! = Quit
|
|
? Menu or option number=28
|
|
Pattern searcher
|
|
? (y/n) (y) Read pattern from keyboard
|
|
X 1 Exact match
|
|
2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =2
|
|
? Motif name=avlst
|
|
? String=avlst
|
|
? Minimum matches (1.00-5.00) (5.00) =3
|
|
Probability of score 3.0000 = 0.394E-02
|
|
1 Exact match
|
|
X 2 Percentage match
|
|
3 Cut-off score and score matrix
|
|
4 Cut-off score and weight matrix
|
|
5 Direct repeat
|
|
6 Membership of set
|
|
7 Pattern complete
|
|
? 0,1,2,3,4,5,6,7 =7
|
|
? (y/n) (y) Save pattern in a file n
|
|
|
|
Pattern description
|
|
|
|
Motif 1 named avlst is of class 2
|
|
which is a match of score 3. to the string
|
|
avlst
|
|
Probability of finding pattern = 0.3941E-02
|
|
Expected number of matches = 0.2030E+01
|
|
? Maximum pattern probability (0.00-1.00) (1.00) =
|
|
? Minimum pattern score (-9999.00-9999.00) (-9999.00) =
|
|
Select display mode
|
|
X 1 Motif by motif
|
|
2 Inclusive
|
|
3 Graphical
|
|
4 EMBL feature table
|
|
? 0,1,2,3,4 =4
|
|
Searching
|
|
|
|
FT avlst 152 156 Program
|
|
Total matches found 1
|
|
Minimum and maximum observed scores 3.00 3.00
|
|
|
|
|
|
General notes
|
|
|
|
These methods allow users to define and search for complex
|
|
patterns of motifs defined as single objects. The programs allow
|
|
individual DNA motifs to be defined in eight different ways, and
|
|
protein motifs in six. Motifs are combined, using the logical
|
|
operators AND, OR and NOT, to describe a pattern. The pattern also
|
|
specifies the ranges of allowed relative separations of the
|
|
individual motifs.
|
|
|
|
First some definitions.
|
|
|
|
A MOTIF is a contiguous subsequence of fixed length. At its
|
|
simplest it could be a single definite base or amino acid; a more
|
|
complex motif might be better represented as a consensus or a weight
|
|
matrix; two more-abstract types of motif are direct and inverted
|
|
repeats.
|
|
|
|
A PATTERN is a higher order of structure defined by a list of
|
|
motifs. The motifs in a pattern are combined using the logical
|
|
operators AND, OR and NOT. The list also defines the allowed
|
|
relative separations of the motifs. In the current versions of the
|
|
programs up to 50 motifs can be combined into a single pattern. So
|
|
using these definitions there are two differences between motifs and
|
|
patterns: 1) the distances between all elements of a motif are
|
|
fixed, but the separations of parts of patterns can vary; 2) all
|
|
characters in a motif are defined using the same method (class), but
|
|
different parts of a pattern can be defined in completely different
|
|
ways.
|
|
|
|
Each motif can be represented in 9 ways (known as the motif
|
|
class):
|
|
|
|
MOTIF CLASSES
|
|
CLASS DESCRIPTION
|
|
1 Exact match to a short defined sequence. The IUB symbols
|
|
can be used for DNA sequences.
|
|
2 Percentage match to a defined short sequence. In nucleic acids,
|
|
the IUB symbols can be used.
|
|
3 Match to a defined sequence, using a score matrix and cutoff
|
|
score. The DNA matrix (see option 18) gives scores to IUB symbols
|
|
depending on their level of redundancy. MDM78 is used for proteins.
|
|
4 Match to a weight matrix with cutoff score.
|
|
5 As class 4 but on the complementary strand.
|
|
6 Inverted repeat or stem-loop. Fixed stem length, range of
|
|
loop sizes, and cutoff score using A-T, G-C=2; G-T=1.
|
|
7 Exact match to short sequence but with a defined step size.
|
|
8 Direct repeat. Fixed repeat length, range of loop-out sizes,
|
|
cutoff score, and score matrix (for protein sequences MDM78 and
|
|
for nucleic acids an identity matrix).
|
|
9 Membership of a set. A list of sets of allowed amino acids for
|
|
each position in the motif. The sets are separated by commas(,).
|
|
For example IVL,,,DEKR,FYWILVM defines a motif of length 5 amino
|
|
acids in which one of I,V or L must be found in the first position,
|
|
then anything in the next two positions, D,E,K or R in the fourth
|
|
position and F,Y,W,I,L,V or M in the fifth. This class only applies
|
|
to protein sequences because for nucleic acids "membership of a
|
|
set"
|
|
can be achieved using IUB symbols.
|
|
|
|
Classes 1 - 4, 8 and 9 apply to protein sequences, and classes 1-8 to
|
|
nucleic acids.
|
|
|
|
|
|
Class 1: exact match.
|
|
|
|
The motif is defined by a short sequence, which for nucleic
|
|
acids, may include IUB symbols. All symbols must match.
|
|
|
|
Class 2: percentage match
|
|
|
|
The motif is defined by a short sequence, which for nucleic
|
|
acids, may include IUB symbols. The minimum number of matching
|
|
characters must also be specified.
|
|
|
|
Class 3: match using a score matrix
|
|
|
|
The motif is defined by a short sequence, which for nucleic
|
|
acids, may include IUB symbols. The motif is not compared directly
|
|
with the sequence to count the number of matching characters.
|
|
Instead a matrix is used to provide a score for all possible pairs
|
|
of characters. The motif score for any position along the sequence
|
|
is the sum of the scores found by looking-up the scores for each
|
|
pair of aligned characters. A match is declared if some minimum
|
|
score is achieved.
|
|
|
|
Class 4: weight matrix
|
|
|
|
The motif is defined by a table of values (called weights or
|
|
scores). The table gives a score for finding each possible character
|
|
at each position along the length of the motif. It therefore has
|
|
dimension motif-length x character-set-size, and allows us to give
|
|
different scores for each character at each position. It is
|
|
equivalent to having a different score matrix for each position
|
|
along the motif, and provides the most flexible and specific method
|
|
of defining motifs. The weight matrices are created by program PIP
|
|
option 20 and stored as files. The file contains the values for each
|
|
position, as well as an overall minimum score. There are two ways in
|
|
which these values can be used to calculate an overall score for any
|
|
section of the sequence. The simplest way is to add the values in
|
|
the file. (This means that the highest possible score can be
|
|
calculated by adding the top value at each column position, and the
|
|
lowest by adding the bottom value.) The normal way of using the
|
|
values in the file is as follows. First the programs divide the
|
|
values in each column by the column total so that they sum to 1.0
|
|
Then the natural logs of these values are used as scores. When the
|
|
matrix is applied to a sequence these logarithmic values are summed
|
|
(which is of course equivalent to multiplying the frequencies).
|
|
Note that using the natural logs of the frequencies as weights and
|
|
adding them means that the overall cutoff score must be less than
|
|
zero, whereas if the original values in the weight matrix file are
|
|
added, the cutoff score will be greater than zero. The search
|
|
routines therefore decide whether the user wants to add values or
|
|
multiply frequencies by examining the value of the cutoff score: it
|
|
will add if the cutoff is greater than zero and add logs of
|
|
frequencies if it is less than zero. Hence we effectively get two
|
|
motif classes in one. The program PIP, when creating weight matrix
|
|
files, will ask the user whether the scores should be added or
|
|
multiplied. If the values in the table have been defined without
|
|
using a set of aligned sequences it is easier for the user to choose
|
|
a cutoff score if the values are added.
|
|
|
|
Class 5: complement of weight matrix
|
|
|
|
The motif is defined by a weight matrix, but the program
|
|
searches for its complement.
|
|
|
|
Class 6: inverted repeat, or stem-loop
|
|
|
|
The motif is defined by a repeat length, a minimum score and a
|
|
range of loop sizes. The scores are A-T=2, G-C=2, G-T=1, else=0.
|
|
The loop sizes are defined by a minimum and maximum distance from
|
|
the 3' end of the stem. For a stem-loop these will be positive
|
|
numbers. For example to define a stem of length 8 and loop sizes
|
|
varying from 3 to 5, the stem would be set to 8, the minimum start
|
|
distance to 3 and the maximum to 5. To define an inverted repeat the
|
|
minimum distance will be negative. For example stem length=9,
|
|
minimum distance=-9, and maximum distance=-8 will find inverted
|
|
repeats of lengths 9 and 10. E.g. AAAAATTTT and AAAAATTTTT would be
|
|
found, the first having a base at its centre, the second having
|
|
none.
|
|
|
|
Class 7: exact match, defined step size.
|
|
|
|
The motif is defined by a short sequence, which for nucleic
|
|
acids, may include IUB symbols. All symbols must match. The class
|
|
differs from class 1 in that searches will move in steps of some
|
|
given size. For example we could search for a certain codon and use
|
|
a step size of 3 and hence keep in a single reading frame.
|
|
|
|
Class 8: direct repeat
|
|
|
|
The motif is defined by a repeat length, a minimum score and a
|
|
range of loop sizes. The scores are defined using MDM78 for protein
|
|
sequences and an identity matrix for nucleic acids. The loop sizes
|
|
are defined by a minimum and maximum distance from the 3' end of the
|
|
stem.
|
|
|
|
Class 9: membership of a set
|
|
|
|
This motif class is for protein sequences. It is defined by
|
|
lists of allowed amino acids for each position in the motif, and a
|
|
cut-off score. Positions at which any amino acid can occur are left
|
|
blank. All allowed amino acids for each position give a score of 1.
|
|
The motifs can be defined in two ways: either typed at the keyboard
|
|
or read in as a weight-matrix-like file. When the motif is defined
|
|
at the keyboard the sets of allowed amino acids are separated by
|
|
commas(,). For example IVL,,,DEKR,FYWILVM defines a motif of length
|
|
5 amino acids in which one of I,V or L must be found in the first
|
|
position, then anything in the next two positions, D,E,K or R in the
|
|
fourth position and F,Y,W,I,L,V or M in the fifth. To specify that
|
|
the whole motif must match a score of 3 would be required (i.e. one
|
|
of the allowed amino acids must be found for each of the three
|
|
defined positions). If the motif is read from a file the file must
|
|
have been written by program PIP, or have been saved by the pattern
|
|
searching routines. If the user elects to save a pattern, and it
|
|
includes class 9 motifs typed at the keyboard, then the program will
|
|
save the class 9 motifs as weight matrix files. Therefore it will
|
|
request file names for each motif of this class. If the motif given
|
|
above as an example were saved the weight matrix file would have 5
|
|
columns. The first column would contain zeroes except for the I, V
|
|
and L rows which would be set to 1; the next two columns would all
|
|
be zero; the next would be zero except for the D,E,K and R rows
|
|
which would be 1; the final column would contain 1's in rows
|
|
F,Y,W,I,L,V and M, with the rest zero.
|
|
|
|
The logical operator (AND, OR or NOT) used to add each motif
|
|
to the pattern is specified by preceding the class number by the
|
|
letters A, O or N. A = AND, O = OR, N = NOT. The default is A, so
|
|
N2 means include, using the NOT operator, a class 2 motif; O2 means
|
|
include, using the OR operator, a class 2 motif; both A2 and 2 mean
|
|
include, using the AND operator, a class 2 motif.
|
|
|
|
Range setting.
|
|
|
|
The motifs in a pattern are numbered according to their order
|
|
in the list. Apart from the first motif in a pattern all motifs are
|
|
given a range of allowed positions relative to a motif further up
|
|
the list. For example suppose we have a pattern defined by A AND B
|
|
AND C AND D. Motif A can occur anywhere, but B must have its range
|
|
of allowed positions defined relative to the position of motif A,
|
|
and C's positions can be defined relative to either A or B,
|
|
depending on which is most convenient, and likewise D's positions
|
|
can be relative to A or B or C.
|
|
|
|
Notice that the positions of motifs can be defined relative to
|
|
more than one motif. Suppose we have a pattern consisting of motifs
|
|
A, B and C, and that B occurs 5-10 residues right of A, C occurs 5-
|
|
10 residues right of B, and also C is never more than 15 residues
|
|
from A. Then it is quite consistent with the methods to include
|
|
motif C into the pattern twice using the AND operator: once relative
|
|
to A and once relative to B. This will define the relative spacing
|
|
and the ORDER of the motifs in the pattern. (If we simply defined
|
|
the position of C relative to A it could be found to the left of B).
|
|
|
|
Motifs combined together using the OR operator are all given
|
|
the same range. For example suppose we had a pattern A AND (B OR C)
|
|
AND (D OR E), then B and C each have the same range, and D and E
|
|
also have the same range as one another. The range for D and E can
|
|
be relative to A or to B.
|
|
|
|
Motifs cannot have their ranges defined relative to motifs
|
|
that are included using the NOT operator. For example if we had the
|
|
pattern A NOT B AND C, then the range for C can only be defined
|
|
relative to motif A.
|
|
|
|
Speed can be gained by arranging the order of the motifs so
|
|
that those higher up the list are of types that can be searched for
|
|
rapidly and that are also unlikely to be found.
|
|
|
|
Motifs combined by the OR operator are alternatives: if any
|
|
one of a set of motifs combined by the OR operator is found, then a
|
|
match is declared. All alternatives will be reported. For example if
|
|
we had a pattern defined by A AND (B OR C), then all places where A
|
|
occurs and B is found within range, and all places where A is found
|
|
and C is found within range will be reported. A typical use would be
|
|
where we might allow a motif to appear on either strand of the DNA
|
|
sequence. For example a weight matrix representing the heatshock
|
|
element could be used in a pattern which included heatshock as a
|
|
motif class 4 combined using the OR operator with heatshock as a
|
|
motif class 5.
|
|
|
|
The probability calculations are performed for each motif as
|
|
it is defined. If an overall probability cut-off is given the
|
|
calculation is repeated for each match found. To achieve maximum
|
|
searching speed do not give an overall probability cut-off. Overall
|
|
cut-off scores should only be used if the motif classes used are
|
|
compatible.
|
|
|
|
There are currently several ways to display the matches: 1 =
|
|
each motif and its position is listed; 2 = all the sequence between
|
|
the two outermost motifs is listed; 3 = graphical, with a spike
|
|
marking the position of the leftmost motif. The library versions
|
|
also give entry names, and a one line title; in addition they can be
|
|
used to produce aligned families of sequences. When this mode of
|
|
output is selected the program will write a separate file for each
|
|
match. The files will be called ENTRYNAME.DAT where ENTRYNAME is the
|
|
name of the entry in the library. The matching sequence will be
|
|
written out so that the spacing between motifs is constant, and set
|
|
to the maximum allowed by the pattern definition. Any gaps will be
|
|
filled with dashes (-). If the individual sequences were
|
|
subsequently written one above the other they should line up so that
|
|
all motifs are in register. There two types of output of this sort:
|
|
one, option 4, writes out whole sequences, the other, option 5,
|
|
writes out only the sequences between the two outermost motifs. If
|
|
the individual sequences were subsequently written one above the
|
|
other they should line up so that all motifs are in register. There
|
|
two types of output of this sort: one, option 4, writes out whole
|
|
sequences, the other, option 5, writes out only the sequences
|
|
between the two outermost motifs. Note that for option 4 users are
|
|
asked to type the position of the first motif, and the reason for
|
|
this is explained below. Consider a pattern found in several
|
|
sequences. Consider only the first motif in the pattern and suppose
|
|
that it was found in different positions in these sequences. Say
|
|
that of these positions the one furthest from the left end was
|
|
position 100. Then, in order to ensure that all the sequences would
|
|
align, we must specify that motif 1 must start at position 100. Any
|
|
sequences in which motif 1 started nearer to the left end than
|
|
position 100 would be padded accordingly. These modes of output
|
|
should only be used when the position of each motif is defined
|
|
relative to its immediate neighbour.
|
|
|
|
The pattern descriptions can be saved to files. These files
|
|
can be used instead of typing definitions again at the keyboard. As
|
|
the files are annotated, they can easily be changed using system
|
|
editors, and the modified versions used to define the variant
|
|
patterns for the programs.
|
|
|
|
|
|
Use of lists of entry names
|
|
|
|
The two programs that operate on libraries have the ability to
|
|
restrict their searches to subsets of the libraries. This does not
|
|
require sublibraries to be created but instead is achieved by using
|
|
files containing a list of the entry names of sequences. The user
|
|
may choose to search only those entries on the list or,
|
|
alternatively to search all but those on the list (i.e. in the
|
|
latter case the list contains the names of those to be excluded).
|
|
The programs can search libraries that have indexes and those that
|
|
do not. If a list of names for inclusion is used, then the search
|
|
will be faster if the index is present. In all other circumstances
|
|
the whole library will be read. The list must be in library order
|
|
except when it is used to include entries, and an index is
|
|
available. The list must contain each entry name on a separate
|
|
line, with the name starting in column 1 of the line. ie there must
|
|
be no spaces at the start of the line. The list of entry names can
|
|
be produced by the keyword searches of nip, pip, etc as long as the
|
|
listings produced have a space character separating the entry name
|
|
from the entry description. This will depend on how well the library
|
|
reformatting programs work. For example swissprot entry names tend
|
|
to run into the beginning of the descriptions, but other libraries
|
|
are generally OK.
|
|
|
|
One use of the programs is to look for patterns that we
|
|
already know about, but in new sequences. However it is hoped that
|
|
they will also be useful for finding new motifs. For example several
|
|
known control regions in nucleic acid sequences consist of
|
|
particular direct or inverted repeats; the inclusion of direct and
|
|
inverted repeats as motif classes makes it possible to find
|
|
previously unknown motifs of these types. Using these new programs
|
|
we can ask questions like: "are there any inverted or direct repeats
|
|
near to sections of sequence that contain both a CCAAT box and a
|
|
TATA box?"; and to search for such things throughout the libraries.
|
|
In addition, the mode of output in which all the sequence between
|
|
the two outermost motifs found is printed out, allows us to extract
|
|
sequences and examine them in more detail for further common
|
|
subsequences. For example we might want to collect together all the
|
|
sequences between putative CCAAT and TATA boxes.
|
|
|
|
A further use of the inverted repeat motif class is the
|
|
following. If a regulatory sequence in DNA is poorly defined but
|
|
also an inverted repeat, then it might be an advantage to specify it
|
|
both as a consensus sequence and a superimposed inverted repeat. In
|
|
this way two weak definitions can be combined to produce a stronger
|
|
pattern.
|
|
|
|
Given only a few examples of a motif it should be possible to
|
|
perform initial searches using a class 3 motif, and then, using
|
|
plausible matching sequences, create a more specific weight matrix
|
|
for the same motif.
|
|
|
|
If motifs are combined with the first motif using the OR
|
|
operator they will be ignored until all permutations that include
|
|
the first motif have been looked for. The whole search will then be
|
|
repeated, in turn, for each of those motifs that are combined with
|
|
the first motif using the OR operator. An interesting consequence
|
|
of this is that the program can be used, without change, to compare
|
|
any newly determined sequence with all known individual motifs. We
|
|
achieve this by having a pattern in which all known relevant motifs
|
|
are combined using the OR operator. If we ask to use this pattern
|
|
with a sequence, the program will automatically compare each
|
|
individual motif in the pattern with the whole length of the
|
|
sequence. As the number of known motifs grows this should become an
|
|
increasingly useful standard procedure.
|
|
|
|
The NOT operator is obviously useful for making sure
|
|
particular motifs are not present, but it can also be used to
|
|
bracket the levels of matches found. We may want a degree of match
|
|
that lies between two limits - binding should occur, but not too
|
|
strongly; or base-pairs should form, but not too many. We can
|
|
specify this by asking for a match with a low score, in combination
|
|
with a match and a high score, both for the same motif, but with the
|
|
high score included using the NOT operator.
|
|
|
|
The algorithm is designed to find all sections of a sequence
|
|
that satisfy the pattern rather than only the best match.
|
|
Particularly if some of the motifs in a pattern are less well
|
|
defined than others, this can often result in the same region of a
|
|
sequence being reported as having several matches, but which only
|
|
vary in the positions of the weakest motifs.
|
|
|
|
General remarks on motif searching
|
|
|
|
Generally motifs are short subsequences that are thought to be
|
|
associated with particular functions in some known sequences. Often
|
|
we search for them to try to understand or interpret other
|
|
sequences. Sometimes we search for motifs and patterns to test a
|
|
hypothesis about their role: are they found in the expected
|
|
positions in the expected sequences. In doing so we should remember
|
|
that, in both proteins and nucleic acids, what we are really looking
|
|
for is a particular three dimensional structure with certain
|
|
affinities for other structures, and that we are assuming that the
|
|
sequence of the motif alone defines the 3D structure we searching
|
|
for. The overall structure may be completely different to those in
|
|
which the motif is functional, and hence the motif may have a
|
|
different shape or be inaccessible. We should be aware of the
|
|
importance of the context in which a motif is found. Where does it
|
|
lie relative to the overall structure, is it accessible, is the
|
|
three dimensional spacing between it and other motifs correct? For
|
|
example, is it on the same side of the double helix, and the correct
|
|
distance from some other motif? How does context affect our
|
|
assessment of the significance of finding a motif? Finding false
|
|
mammalian mRNA splice junctions in non-coding sequences is far less
|
|
important than finding false sites in pre-mRNA sequences, but
|
|
finding them in the correct places is most important! In other
|
|
words, it is often the case that when we are searching for a motif
|
|
that is known to be necessary for some function, then a positive
|
|
result in the form of a match in the required position, is more
|
|
important than a high background of matches in the wrong positions.
|
|
Being able to write down the probability of finding a motif in a
|
|
random sequence tells us how well it is defined. In nucleic acids
|
|
the DNA may contain many superimposed types of information such as
|
|
those concerned with histone phasing, protein coding or mRNA
|
|
secondary structure. These overlapping "codes" may interfere with
|
|
one another causing matches to motifs to be poorer than expected.
|
|
In general we will only have a limited number of examples of the
|
|
motif and we do not know how representative they are.
|
|
|
|
Sequences have superimposed functions: some parts may be of
|
|
general structural importance and give rise to an overall framework,
|
|
and other parts give specificity and hence are not common; we may
|
|
want to use a set of aligned sequences to define a motif, but want
|
|
to use only the framework positions. Alternatively we may want to
|
|
pick out only those parts of a set of aligned sequences that give a
|
|
particular property, and to ignore other similarities that are due
|
|
to some other property and which could obscure the pattern we are
|
|
interested in. It is possible to apply a mask to a set of aligned
|
|
sequences in order to give weight to selected positions only. The
|
|
ability to define a mask allows certain positions to be used in the
|
|
motif and others to be ignored, and yet still permits the use of a
|
|
set of aligned sequences to calculate weights. The mask is requested
|
|
and applied by the program and results in the masked positions being
|
|
zero in the weight matrix. The mask is defined in the following way.
|
|
Suppose we had a motif of length 15, then the mask x--x--xx-x will
|
|
give zero weights to positions 2,3,5,6 and 9 (note it is the dashes
|
|
(-) that are significant and that positions 1,4,7,8,10,11,12,13,14
|
|
and 15 will be non-zero). Of course the same set of sequences could
|
|
be used with several alternative masks in order to extract different
|
|
features and create corresponding weight matrices.
|
|
|
|
The programs are described in Staden,R. CABIOS 4, 53-60, 1988;
|
|
Staden,R. CABIOS 5, 89-96, 1989, anf a forthcoming Methods in
|
|
Enzymology.
|
|
@ end of help
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