STEMLOOP

Table of Contents

FUNCTION

DESCRIPTION

EXAMPLE

FUNCTION

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StemLoop finds stems (inverted repeats) within a sequence. You specify the minimum stem length, minimum and maximum loop sizes, and the minimum number of bonds per stem. All stems or only the best stems can be displayed on your screen or written into a file.

DESCRIPTION

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StemLoop searches for inverted repeats in your sequence after you choose a minimum stem length and minimum and maximum loop sizes. You must also specify a minimum number of bonds per stem with G-T, A-T/U, and G-C scored as 1, 2, and 3 bonds, respectively. The stems found can be sorted by position, size (stem length), or quality (number of bonds) and can be either filed or displayed on the screen. StemLoop tells you the number of stems found for your settings of minimum stem size, maximum loop size, minimum loop size, and minimum bonds per stem. If you feel there are too many stems, you may reset the parameters without reviewing the stems found or view only the best stems found. To view only the best stems, there must be more than 25 stems found and you must sort them by quality or size. (See the ALGORITHM topic below to understand precisely what StemLoop does.)

EXAMPLE

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Here is a session using StemLoop to see the 8 inverted repeats that have at least 18 bonds within stems at least 8 base pairs in length in an Alu consensus sequence:

% stemloop

  STEMLOOP of what sequence ?  alucons.seq

                   Begin (* 1 *) ?

                 End (*   290 *) ?

  What minimum stem length (* 6 *) ?  8

  What minimum number of bonds/stem (* 16 *) ?  18

  What maximum loop size (* 20 *) ?

  What minimum loop size (* 3 *) ?

              just a second  ...

  There are 8 stems.  Would you like to:

        1) See the stems

        2) See the stem coordinates

        3) File the stems

        4) File the stems as points for DOTPLOT

        5) Choose new parameters,

        6) Get a different sequence

        Q)uit ?

  Please choose one (* 1 *):  3

  Sort Stems by:

        1) Position

        2) Quality

        3) Size

        Q)uit

  Please choose one (* 1 *):  2

  What should I call the output file (* alucons.stem *) ?

  There are 8 stems.  Would you like to:

        1) See the stems,

        2) See the stem coordinates,

        3) File the stems,

        4) File the stems as points for DOTPLOT

        4) Choose new parameters,

        5) Get a different sequence

        Q)uit ?

  Please choose one (* Q *):

OUTPUT

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StemLoop creates an output file if you choose to file the stems from any search; otherwise, you may view the stems on your screen. In either case, the stem is shown, as below, with vertical bars ('|') indicating the base pairs. The associated loop is shown to the right of the stem. If either the stem or loop is too long to be displayed in its entirety on the line, then only that part that fits on the line is shown. The first and last coordinates of the stem are displayed on the left, and the length of the stem (size), the number of bonds in the stem (quality), and the loop size are shown on the right. Here is part of the file alucons.stem created by the example session above:

  STEMLOOP of: alucons.seq  check: 1861  from: 1  to: 290

Alu consensus sequence

Labuda, D. and Striker, G. (1989) Sequence conservation in Alu

evolution.  Nucleic Acids Research 17, 2477-2491.

 Minimum Stem: 8  Minimum bonds/stem: 18  Maximum loop size: 20

 Stems found: 8  Stems shown: 8

 Average Match: 1.80  Average Mismatch: 0.00  Nibbling Threshold:  1

                           October 6, 1998 14:16  ..

    217 AGGCTGCAGTG  AGCCGTGAT    11, 25

        |||||| ||||           C

    257 TCCGGCCTCAC  GTCACCGCG    19

    135 TAGCCGGGCGT  GG    11, 22

        ||| || ||||

    160 GTCCGCGCGCG  GT    4

    /////////////////////////////

    221 TGCAGTG  AGCCGTG    7, 18

        |||||||

    248 ACGTCAC  CGCGCTA    14

     35 CACTTCGG  GA    8, 18

        | ||||||

     54 GCGGAGCC  GG    4

You may choose to see only the numbers defining each stem on your screen by choosing option '2' in the first menu. This is what that screen output would look like if you choose option '2' in the first menu and then choose to sort by quality in the second menu:

        Loop     Start       End      Size     Quality

           1       217       257        11          25

           2       135       160        11          22

           3       139       160         8          20

           4        69        95         7          20

           5         4        25         9          20

           6       213       247         8          19

           7       221       248         7          18

           8        35        54         8          18

StemLoop can also make an output file with points for plotting with DotPlot.

INPUT FILES

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StemLoop accepts a single nucleotide sequence as input. If StemLoop rejects your nucleotide sequence, turn to Appendix VI to see how to change or set the type of a sequence.

RELATED PROGRAMS

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MFold predicts optimal and suboptimal secondary structures for an RNA or DNA molecule using the most recent energy minimization method of Zuker. PlotFold displays the optimal and suboptimal secondary structures for an RNA or DNA molecule predicted by MFold.

Using Compare-DotPlot to create a dot-plot of the similarities between a nucleotide sequence and its reverse-complement strand is functionally equivalent to running StemLoop. Repeat uses the same algorithm as StemLoop to find repeats that are not inverted. DotPlot shows you the output from Compare or StemLoop on a surface of comparison.

RESTRICTIONS

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StemLoop only searches for loops through a range that is equal to twice the minimum stem length, plus the maximum loop size. You may extend the search range by increasing the maximum loop size; however, the maximum range for the search may not exceed 2,000 bases. StemLoop cannot find more than 1,000 loops.

ALGORITHM

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StemLoop uses a window and stringency match criterion in exactly the same manner as Compare. For every position in each register shift, a window set by you as the minimum stem size is moved along the sequence, and if the minimum number of bonds per stem or more are found, then a stem is recorded covering all of the bases under the window. The number of the bonds under the window at each window position is the sum of the scoring matrix values for each base pair found in the file stemloop.cmp (see the LOCAL DATA FILES topic below) . Mismatches can be scored negatively, although the public data file simply scores matches with G-T, A-T/U, and G-C worth 1, 2, and 3, respectively. Several adjacent mismatches may be found within a long stem if there are strong matches on either side. The criterion for a stem is that the minimum number of bonds occur within a length set by you as the minimum stem length.

Stem Extension and Nibbling

Before the stems are presented, they are extended (or nibbled) from both ends so that the first base on each end participates in a bond. The criterion for a bond between pairing bases is that the value in the scoring matrix file (stemloop.cmp) for the pair is greater than or equal to the average positive non-identical comparison value in the scoring matrix. You can reset the threshold for nibbling with -PAIr. You could set a pairing threshold high enough so that all stems are nibbled away!

Since stem nibbling occurs, stems shorter than the minimum stem length are commonly reported. If, on the other hand, extra pairing bases are found adjacent to the stem, the stem is extended until a pair of bases do not have a bond between them. If the nibbling function finds, after nibbling a stem, that the now enlarged loop is longer than the specified maximum loop size, the stem is not reported.

CONSIDERATIONS

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StemLoop chooses a default minimum number of bonds per stem that is appropriate for the scoring matrix it reads. If you select a different scoring matrix with -MATRix, the program will adjust the default minimum number of bonds per stem accordingly.

COMMAND-LINE SUMMARY

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All parameters for this program may be added to the command line. Use -CHEck to view the summary below and to specify parameters before the program executes. In the summary below, the capitalized letters in the parameter names are the letters that you must type in order to use the parameter. Square brackets ([ and ]) enclose parameter values that are optional.

Minimal Syntax: % stemloop [-INfile=]alucons.seq -Default

Prompted Parameters:

-BEGin=1 -END=290           sets the range of interest

-STEMlength=6               sets the minimum stem length

-BONds=12                   sets the minimum bonds per stem

-MINLoopsize=3              sets the minimum loop size

-MAXLoopsize=20             sets the maximum loop size (distance to furthest

                              inverted repeat)

-MENu1=1                    specifies output type: 1=see stems,

                              2=see coordinates, 3=file, 4=dotplot file

-MENu2=1                    sorts by: 1=position, 2=quality, 3=size

-MAXSTems=25                sets the maximum number of stems to show

                              (quality or size sorts only)

[-OUTfile=]alucons.stem     names the output file

Local Data Files:

-MATRix=stemloop.cmp        assigns the scoring matrix for finding bonds/stem

Optional Parameters:

-PAIr=1                     sets threshold for nibbling, match (|),

                              and point display

Note: StemLoop does not cycle through the menus repeatedly if you specify either -MENu1 or -MENu2 on the command line.

LOCAL DATA FILES

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

Local Scoring Matrices

This program reads one or more scoring matrices for the comparison of sequence characters. The program automatically reads the program's default scoring matrix in a public data directory unless you either 1) have a data file with exactly the same name as the program default scoring matrix in your current working directory; or 2) have a data file with exactly the same name as the program default scoring matrix in the directory with the logical name MyData; or 3) name a file on the command line with an expression like -MATRix=mymatrix.cmp. If you don't include a directory specification when you name a file with -MATRix, the program searches for the file first in your local directory, then in the directory with the logical name MyData, then in the public data directory with the logical name GenMoreData, and finally in the public data directory with the logical name GenRunData. For more information see "Using a Special Kind of Data File: A Scoring Matrix" in Section 4, Using Data Files in the User's Guide.

StemLoop uses a scoring matrix of the kind described in Appendix VII to find the number of bonds between any possible pair of bases. Every non-zero value is defined in the scoring matrix. StemLoop reads the scoring matrix file stemloop.cmp in your local directory, or if it fails to find such a file there, it uses the public file of the same name. The file can be customized so that any score, positive or negative, can be assigned to any possible pair of bases (GCG symbols). You can get the public file with % fetch stemloop.cmp. The values in the file assign G-T, A-T/U, and G-C to 1, 2, and 3 respectively, with all other pairs valued at zero. A more realistic set of values might assign some negative score to the mismatches, especially purine-purine pairs. This would make the output sorted by quality more significant.

PARAMETER REFERENCE

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You can set the parameters listed below from the command line.

-STEMlength=6

Sets the minimum stem length.

-BONds=12

Sets the minimum bonds per stem.

-MINLoopsize=3

Sets the minimum loop size.

-MAXLoopsize=20

Sets the maximum loop size (distance to furthest inverted repeat).

-MENu1=1

Indicates the type of output. -MENu1=1 means display the stems; other values for -MENu1 are: 2) display the coordinates, 3) save the stems to a file, and 4) save the stem coordinates to a DotPlot file.

-MENu2=1

Indicates how to sort the stems in the output. -MENu2=1 means sort by position; other values for -MENu2 are: 2) sort by quality and 3) sort by size.

-MAXSTems=25

Sets the maximum number of stems to show (only applies when stems are sorted by quality or size).

-MATRix=mymatrix.cmp

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

For more information see the Local Scoring Matrices section.

-PAIr=1

The output from this program has a '|' (vertical bar) between sequence symbols that match. This match display character is added to the output whenever the symbol comparison value for the two symbols in your scoring matrix is greater than or equal to the average positive non-identical comparison value in the matrix. The -PAIr parameter lets you specify a match display threshold appropriate for the scoring matrix you are using.

Stem structure nibbling also uses the threshold value set by this parameter to decide what pairs should be nibbled away from the structure. You can set a pairing threshold high enough so that all stems are nibbled away!

Printed: May 27, 2005 14:47

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