EMBOSS

| ## EMBOSS: vectorstrip

# Program vectorstrip

## Function

Strips out DNA between a pair of vector sequences

## Description

vectorstrip is intended to be useful for stripping vector sequence from the ends of sequences of interest. For example, if a fragment has been cloned into a vector and then sequenced, the sequence may contain vector data eg from the cloning polylinker at the 5' and 3' ends of the sequence. vectorstrip will remove these contaminating regions and output trimmed sequence ready for input into another application.

vectorstrip is suitable for use with low quality sequence data as it can allow for mismatches between the sequence and the vector patterns provided. You can specify the maximum level of mismatch expected.

Vector data can either be provided in a file or interactively. If presented in a file, vectorstrip will search all input sequences with all vectors listed in that file. The intention is that the user can maintain a single file for use with vectorstrip, containing all the linker sequences commonly used in the laboratory.

The two patterns for each vector are searched separately against the sequence. Once the search is completed, each of the hits of the 5' sequence is paired with each of the hits of the 3' sequence and the resulting subsequences are output. For example, if the 5' sequence matches the sequence from (a) position 30-60, and(b)position 70-100, and the 3' sequence matches from 150-175, then two subsequences will be output: from 61-149, and from 101-149. The lower the quality of the sequence, the more likely multiple hits become if nonzero mismatches are accepted.

Default behaviour is to report only the best matches between the vector patterns and the sequence. This means that if you specify a maximum mismatch level of 10%, but the vector patterns match the sequence with zero mismatches, the search will stop and the program will output only these "best" matches. If there are no perfect matches, the program will try searching again allowing 1 mismatch, then 2, and so on until either the patterns match the sequence or the maximum specified mismatch level is exceeded. You can tell vectorstrip to show all possible matches up to your specified maximum level, as illustrated in the examples below.

## Usage

Here are several examples of running vectorstrip. The vectorfile and the sequence files used in these example are given below. In each case, the same fragment has been cloned into the XhoI site of the polylinker of each vector. The cloned fragment is represented in lower case and the vector sequence in upper case so the sequence trimming can be readily seen.

1. Running vectorstrip on a list of sequences with default parameters

       unix % vectorstrip @seqs.list
       Strips out DNA between a pair of vector sequences
       Are your vector sequences in a file? [Y]:
       Name of vectorfile: vectors
       Max allowed % mismatch [10]:
       Show only the best hits (minimise mismatches)? [Y]:
       Output file [pbluescript.vectorstrip]:stdout
       Output sequence [pbluescript.fasta]:

       Sequence: pBlueScript    Vector: pTYB1  No match

       Sequence: pBlueScript    Vector: pBS_KS+
       5' sequence matches:
               From 67 to 83 with 0 mismatches
       3' sequence matches:
               From 205 to 219 with 0 mismatches
       Sequences output to file:
               from 84 to 204
                       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagca
                       cacagtgacatgagagacacagatatagagacagatagacgatagacaga
                       cagcatatatagacagatagc
               sequence trimmed from 5' end:
                       GGAAACAGCTAATGACCATGATTACGCCAAGCGCGCAATTAACCCTCACT
                       AAAGGGAACAAAAGCTGGGTACCGGGCCCCCCC
               sequence trimmed from 3' end:
                       TCGAGGTCGACGGTATCGATAAGCTTGATATCG

       Sequence: pBlueScript    Vector: pLITMUS        No match

       Sequence: litmus.seq     Vector: pTYB1  No match

       Sequence: litmus.seq     Vector: pBS_KS+        No match

       Sequence: litmus.seq     Vector: pLITMUS
       5' sequence matches:
               From 43 to 61 with 0 mismatches
       3' sequence matches:
               From 183 to 199 with 0 mismatches
       Sequences output to file:
               from 62 to 182
                       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagca
                       cacagtgacatgagagacacagatatagagacagatagacgatagacaga
                       cagcatatatagacagatagc
               sequence trimmed from 5' end:
                       TCTAGAACCGGTGACGTCTCCCATGGTGAAGCTTGGATCCACGATATCCT
                       GCAGGAATTCC

       Sequence: pTYB1.seq      Vector: pTYB1
       5' sequence matches:
               From 40 to 58 with 0 mismatches
       3' sequence matches:
               From 180 to 196 with 0 mismatches
       Sequences output to file:
               from 59 to 179
                       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagca
                       cacagtgacatgagagacacagatatagagacagatagacgatagacaga
                       cagcatatatagacagatagc
               sequence trimmed from 5' end:
                       CTTTAAGAAGGAGATATACATATGGCTAGCTCGCGAGTCGACGGCGGCCG
                       CGAATTCC
               sequence trimmed from 3' end:
                       TCGAGGGCTCTTCCTGCTTTGCCAAGGGTACCAATGTTTTAATGGCGGAT

       Sequence: pTYB1.seq      Vector: pBS_KS+        No match

       Sequence: pTYB1.seq      Vector: pLITMUS        No match

       unix % more pbluescript.fasta
       >pBlueScript_from_84_to_204 KS+
       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagcacacagtgaca
       tgagagacacagatatagagacagatagacgatagacagacagcatatatagacagatag
       c
       >litmus.seq_from_62_to_182
       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagcacacagtgaca
       tgagagacacagatatagagacagatagacgatagacagacagcatatatagacagatag
       c
       >pTYB1.seq_from_59_to_179
       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagcacacagtgaca
       tgagagacacagatatagagacagatagacgatagacagacagcatatatagacagatag
       c

2. Running vectorstrip allowing maximum 30% mismatch, and asking only for best hits

       unix % vectorstrip litmus.seq
       Strips out DNA between a pair of vector sequences
       Are your vector sequences in a file? [Y]:
       Name of vectorfile: vectors
       Max allowed % mismatch [10]: 30
       Show only the best hits (minimise mismatches)? [Y]:
       Output file [litmus.vectorstrip]: stdout
       Output sequence [litmus.fasta]:

       Sequence: litmus.seq     Vector: pTYB1  No match

       Sequence: litmus.seq     Vector: pBS_KS+        No match

       Sequence: litmus.seq     Vector: pLITMUS
       5' sequence matches:
               From 43 to 61 with 0 mismatches
       3' sequence matches:
               From 183 to 199 with 0 mismatches
       Sequences output to file:
               from 62 to 182
                       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagca
                       cacagtgacatgagagacacagatatagagacagatagacgatagacaga
                       cagcatatatagacagatagc
               sequence trimmed from 5' end:
                       TCTAGAACCGGTGACGTCTCCCATGGTGAAGCTTGGATCCACGATATCCT
                       GCAGGAATTCC
               sequence trimmed from 3' end:
                       TCGAGACCGTACGTGCGCGCGAATGCATCCAGATCTTCCCTCTAGTCAAG
                       GCCTTAAGTGAGTCGTATTACGGA

3. Running vectorstrip allowing maximum 30% mismatch, and asking for all hits

       unix % vectorstrip litmus.seq
       Strips out DNA between a pair of vector sequences
       Are your vector sequences in a file? [Y]:
       Name of vectorfile: vectors
       Max allowed % mismatch [10]: 30
       Show only the best hits (minimise mismatches)? [Y]: N
       Output file [litmus.vectorstrip]: stdout
       Output sequence [litmus.fasta]:

       Sequence: litmus.seq     Vector: pTYB1  No match

       Sequence: litmus.seq     Vector: pBS_KS+        No match

       Sequence: litmus.seq     Vector: pLITMUS
       5' sequence matches:
               From 43 to 61 with 0 mismatches
       3' sequence matches:
               From 183 to 199 with 0 mismatches
               From 228 to 244 with 5 mismatches
       Sequences output to file:
               from 62 to 182
                       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagca
                       cacagtgacatgagagacacagatatagagacagatagacgatagacaga
                       cagcatatatagacagatagc
               sequence trimmed from 5' end:
                       TCTAGAACCGGTGACGTCTCCCATGGTGAAGCTTGGATCCACGATATCCT
                       GCAGGAATTCC
               sequence trimmed from 3' end:
                       TCGAGACCGTACGTGCGCGCGAATGCATCCAGATCTTCCCTCTAGTCAAG
                       GCCTTAAGTGAGTCGTATTACGGA

               from 62 to 227
                       tcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagca
                       cacagtgacatgagagacacagatatagagacagatagacgatagacaga
                       cagcatatatagacagatagcTCGAGACCGTACGTGCGCGCGAATGCATC
                       CAGATCTTCCCTCTAG
               sequence trimmed from 5' end:
                       TCTAGAACCGGTGACGTCTCCCATGGTGAAGCTTGGATCCACGATATCCT
                       GCAGGAATTCC
               sequence trimmed from 3' end:
                       TCAAGGCCTTAAGTGAGTCGTATTACGGA

4. Running vectorstrip against a sequence containing Ns

       unix % vectorstrip pTYB1_N.seq
       Strips out DNA between a pair of vector sequences
       Are your vector sequences in a file? [Y]:
       Name of vectorfile: vectors
       Max allowed % mismatch [10]: 30
       Show only the best hits (minimise mismatches)? [Y]:
       Output file [ptyb1.vectorstrip]: stdout
       Output sequence [ptyb1.fasta]:

       Sequence: pTYB1.seq      Vector: pTYB1
       5' sequence matches:
               From 40 to 58 with 2 mismatches
       3' sequence matches:
               From 180 to 196 with 2 mismatches
       Sequences output to file:
               from 59 to 179
                       tcnagagccgtatngcgatatngcgcacatgcgntggacacagangagca
                       cacagtnacatgagagncacagatntagagacagatngacgataganaga
                       cagcatanatagacanatagc
               sequence trimmed from 5' end:
                       CTTTAAGNAGGAGANATACANATGGCNAGCTCGCGANTCGACGGCGGNCG
                       CGAATNCC
               sequence trimmed from 3' end:
                       TCGNGGGCTCTTCCNGCTTTGCCANGGGTACCAANGTTTTAATGGCNGAT

       Sequence: pTYB1.seq      Vector: pBS_KS+        No match

       Sequence: pTYB1.seq      Vector: pLITMUS        No match

## Command line arguments

       Mandatory qualifiers (* if not always prompted):
      [-sequence]          seqall     Sequence database USA
      [-[no]vectorfile]    bool       Are your vector sequences in a file?
    * [-vectors]           infile     Name of vectorfile
       -mismatch           integer    Max allowed % mismatch
       -[no]besthits       bool       Show only the best hits (minimise
                                      mismatches)?
    *  -linkera            string     5' sequence
    *  -linkerb            string     3' sequence
      [-outf]              outfile    Output file name
      [-outseq]            seqoutall  Output sequence(s) USA

       Optional qualifiers: (none)
       Advanced qualifiers: (none)
       General qualifiers:
      -help                bool       report command line options. More
                                      information on associated and general
                                      qualifiers can be found with -help -verbose

Mandatory qualifiers | Allowed values | Default
--- | --- | ---
[-sequence] (Parameter 1) | Sequence database USA | Readable sequence(s) | Required
[-[no]vectorfile] (Parameter 2) | Are your vector sequences in a file? | Yes/No | Yes
[-vectors] (Parameter 3) | Name of vectorfile | Input file | Required
-mismatch | Max allowed % mismatch | Any integer value | 10
-[no]besthits | Show only the best hits (minimise mismatches)? | Yes/No | Yes
-linkera | 5' sequence | Any string is accepted | An empty string is accepted
-linkerb | 3' sequence | Any string is accepted | An empty string is accepted
[-outf] (Parameter 4) | Output file name | Output file | <sequence>.vectorstrip
[-outseq] (Parameter 5) | Output sequence(s) USA | Writeable sequence(s) | <sequence>.format
Optional qualifiers | Allowed values | Default
(none)
Advanced qualifiers | Allowed values | Default
(none)

## Input file format

Each line should contain the name of the vector, the 5' pattern and the 3' pattern.
Lines beginning with # are treated as comments and ignored.
If only one vector sequence is given in the it will be assumed that this is the 5' pattern.
If a vector name is given but no pattern data, the vector will not be used.

For example, the file used in the examples above is:

    pTYB1	GACGGCGGCCGCGAATTCC	TCGAGGGCTCTTCCTGC
    pBS_KS+	GGGTACCGGGCCCCCCC	TCGAGGTCGACGGTA
    pLITMUS	GATATCCTGCAGGAATTCC	TCGAGACCGTACGTGCG

The sequence files used in the examples above are:

1. seqs.list

       bluescript.seq
       litmus.seq
       pTYB1.seq

2. bluescript.seq

       > pBlueScript KS+
       GGAAACAGCTAATGACCATGATTACGCCAAGCGCGCAATTAACCCTCACTAAAGGGAAC
       AAAAGCTGGGTACCGGGCCCCCCCtcgagagccgtattgcgatatagcgcacatgcgtt
       ggacacagatgagcacacagtgacatgagagacacagatatagagacagatagacgata
       gacagacagcatatatagacagatagcTCGAGGTCGACGGTATCGATAAGCTTGATATCG

3. litmus.seq

       > litmus.seq
       TCTAGAACCGGTGACGTCTCCCATGGTGAAGCTTGGATCCACGATATCCTGCAGGAATT
       CCtcgagagccgtattgcgatatagcgcacatgcgttggacacagatgagcacacagtg
       acatgagagacacagatatagagacagatagacgatagacagacagcatatatagacag
       atagcTCGAGACCGTACGTGCGCGCGAATGCATCCAGATCTTCCCTCTAGTCAAGGCCT
       TAAGTGAGTCGTATTACGGA

4. pTYB1.seq

       >pTYB1.seq
       CTTTAAGAAGGAGATATACATATGGCTAGCTCGCGAGTCGACGGCGGCCGCGAATTCCtc
       gagagccgtattgcgatatagcgcacatgcgttggacacagatgagcacacagtgacatg
       agagacacagatatagagacagatagacgatagacagacagcatatatagacagatagcT
       CGAGGGCTCTTCCTGCTTTGCCAAGGGTACCAATGTTTTAATGGCGGAT

5. pTYB1_N.seq

       >pTYB1.seq lower quality
       CTTTAAGNAGGAGANATACANATGGCNAGCTCGCGANTCGACGGCGGNCGCGAATNCCtc
       nagagccgtatngcgatatngcgcacatgcgntggacacagangagcacacagtnacatg
       agagncacagatntagagacagatngacgataganagacagcatanatagacanatagcT
       CGNGGGCTCTTCCNGCTTTGCCANGGGTACCAANGTTTTAATGGCNGAT

In each case the vector sequence is given in upper case and the cloned sequence in lower case.

## Output file format

Two types of output file are produced:

1. The sequence file(s) - contain the trimmed subsequence(s) produced by vectorstrip either all in one file, or in separate files if the command line flag -ossingle is used.
2. Results summary file eg

       Sequence: pTYB1.seq      Vector: pTYB1
       5' sequence matches:
               From 40 to 58 with 2 mismatches
       3' sequence matches:
               From 180 to 196 with 2 mismatches
       Sequences output to file:
               from 59 to 179
                       tcnagagccgtatngcgatatngcgcacatgcgntggacacagangagca
                       cacagtnacatgagagncacagatntagagacagatngacgataganaga
                       cagcatanatagacanatagc
               sequence trimmed from 5' end:
                       CTTTAAGNAGGAGANATACANATGGCNAGCTCGCGANTCGACGGCGGNCG
                       CGAATNCC
               sequence trimmed from 3' end:
                       TCGNGGGCTCTTCCNGCTTTGCCANGGGTACCAANGTTTTAATGGCNGAT

## Data files

None.

## Notes

None.

## References

None.

## Warnings

None.

## Diagnostic Error Messages

1.     No suitable vectors found - exiting

   indicates that the 5' and 3' patterns for the vectors were blank - usually this is as a result of an empty vectorfile.

2.     Illegal pattern

   indicates that one of the vector patterns could not be compiled and therefore cannot be searched.

3.     5' and 3' sequence matches are identical; inconclusive

   indicates that the 5' and 3' patterns provided were identical, and that they only match the sequence once. Thus the program cannot determine which part of the sequence is vector and which is insert.

## Exit status

It always exits with status 0.

## Known bugs

None.

## See also

Program name | Description
--- | ---
biosed | Replace or delete sequence sections
cutseq | Removes a specified section from a sequence
degapseq | Removes gap characters from sequences
descseq | Alter the name or description of a sequence
entret | Reads and writes (returns) flatfile entries
extractfeat | Extract features from a sequence
extractseq | Extract regions from a sequence
listor | Writes a list file of the logical OR of two sets of sequences
maskfeat | Mask off features of a sequence
maskseq | Mask off regions of a sequence
newseq | Type in a short new sequence
noreturn | Removes carriage return from ASCII files
notseq | Excludes a set of sequences and writes out the remaining ones
nthseq | Writes one sequence from a multiple set of sequences
pasteseq | Insert one sequence into another
revseq | Reverse and complement a sequence
seqret | Reads and writes (returns) sequences
seqretsplit | Reads and writes (returns) sequences in individual files
splitter | Split a sequence into (overlapping) smaller sequences
swissparse | Retrieves sequences from swissprot using keyword search
trimest | Trim poly-A tails off EST sequences
trimseq | Trim ambiguous bits off the ends of sequences
union | Reads sequence fragments and builds one sequence
yank | Reads a sequence range, appends the full USA to a list file

## Author(s)

This application was written by Val Curwen (vcurwen@hgmp.mrc.ac.uk)

## History

16 August 2000 - Val Curwen - Written.

## Target users

This program is intended to be used by everyone and everything, from naive users to embedded scripts.

## Comments