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EMBOSS: primersearch |
The user can specify a maximum percent mismatch level; for example, 10% mismatch on a primer of length 20bp means that the program will classify a primer as matching a sequence if 18 of the 20 base pairs matches.
It will only report matches if both primers in the pair have a match in opposite orientations.
unix% primersearch embl:Z52466 Searches DNA sequences for matches with primer pairs Primer file: primers Output file [hsa203yc1.primersearch]: stdout Allowed percent mismatch [0]: Primer name D1S243 Primer name D1S468 Primer name D1S2845 Primer name D1S1608 Primer name D1S2893 Primer name D1S2660 Amplimer 1 Sequence: HSA203YC1 Z52466 H.sapiens (D1S2660) DNA segment containing (CA) repeat; clone AFMa203yc1; single read. CACACATGCACATGCAC hits forward strand at 27 with 0 mismatches AGTGACACCAGCAGGG hits reverse strand at [103] with 0 mismatches Amplimer length: 261 bp
Here we run the same example but allowing 20% mismatch between the primers and the sequence.
unix % primersearch embl:Z52466 Searches DNA sequences for matches with primer pairs Primer file: primers Output file [hsa203yc1.primersearch]: stdout Allowed percent mismatch [0]: 20 Primer name D1S243 Primer name D1S468 Primer name D1S2845 Primer name D1S1608 Primer name D1S2893 Primer name D1S2660 Amplimer 1 Sequence: HSA203YC1 Z52466 H.sapiens (D1S2660) DNA segment containing (CA) repeat; clone AFMa203yc1; single read. CACACATGCACATGCAC hits forward strand at 49 with 2 mismatches AGTGACACCAGCAGGG hits reverse strand at [103] with 0 mismatches Amplimer length: 239 bp Amplimer 2 Sequence: HSA203YC1 Z52466 H.sapiens (D1S2660) DNA segment containing (CA) repeat; clone AFMa203yc1; single read. CACACATGCACATGCAC hits forward strand at 27 with 0 mismatches AGTGACACCAGCAGGG hits reverse strand at [103] with 0 mismatches Amplimer length: 261 bp
Here is an example of running with a file containing a list of sequences.
curium % primersearch @seqs.list Searches DNA sequences for matches with primer pairs Primer file: primers Output file [hs214yg7.primersearch]: stdout Allowed percent mismatch [0]: Primer name D1S243 Amplimer 1 Sequence: HS214YG7 Z16979 H. sapiens (D1S243) DNA segment containing (CA) repeat; clone AFM214yg7; single read. CACACAGGCTCACATGCC hits forward strand at 122 with 0 mismatches GCTCCAGCGTCATGGACT hits reverse strand at [36] with 0 mismatches Amplimer length: 162 bp Primer name D1S468 Amplimer 1 Sequence: HS280WE5 Z23994 H. sapiens (D1S468) DNA segment containing (CA) repeat; clone AFM280we5; single read. AATTAACCGTTTTGGTCCT hits forward strand at 47 with 0 mismatches GCGACACACACTTCCC hits reverse strand at [96] with 0 mismatches Amplimer length: 185 bp Primer name D1S2845 Amplimer 1 Sequence: HS344WE9 Z51474 H.sapiens (D1S2845) DNA segment containing (CA) repeat; clone AFM344we9; single read. CCAAAGGGTGCTTCTC hits forward strand at 29 with 0 mismatches GTGGCATTCCAACCTC hits reverse strand at [157] with 0 mismatches Amplimer length: 201 bp Primer name D1S1608 Amplimer 1 Sequence: HS829186 G07829 human STS CHLC.GATA49A06.P15262 clone GATA49A06, sequence tagged site. GATGGCTTTTGGGGACTATT hits forward strand at 13 with 0 mismatches CACTGAGCCAAGTGACACAG hits reverse strand at [92] with 0 mismatches Amplimer length: 270 bp Primer name D1S2893 Amplimer 1 Sequence: HS123XC3 Z50993 H.sapiens (D1S2893) DNA segment containing (CA) repeat; clone AFM123xc3; single read. AAAACATCAACTCTCCCCTG hits forward strand at 5 with 0 mismatches CTCAAACCCCAATAAGCCTT hits reverse strand at [3] with 0 mismatches Amplimer length: 215 bp Primer name D1S2660 Amplimer 1 Sequence: HSA203YC1 Z52466 H.sapiens (D1S2660) DNA segment containing (CA) repeat; clone AFMa203yc1; single read. CACACATGCACATGCAC hits forward strand at 27 with 0 mismatches AGTGACACCAGCAGGG hits reverse strand at [103] with 0 mismatches Amplimer length: 261 bp
Mandatory qualifiers: [-sequences] seqall Sequence database USA [-primers] infile Primer file [-mismatchpercent] integer Allowed percent mismatch [-out] outfile Output file name 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 | |
---|---|---|---|
[-sequences] (Parameter 1) |
Sequence database USA | Readable sequence(s) | Required |
[-primers] (Parameter 2) |
Primer file | Input file | Required |
[-mismatchpercent] (Parameter 3) |
Allowed percent mismatch | Any integer value | 0 |
[-out] (Parameter 4) |
Output file name | Output file | <sequence>.primersearch |
Optional qualifiers | Allowed values | Default | |
(none) | |||
Advanced qualifiers | Allowed values | Default | |
(none) |
Comment lines start with a '#'
Lines with primer information have three fields separated by spaces or TAB characters. The columns contain:
Empty files will cause primersearch to note that no primers have been found, and to exit.
An example of the primer file follows:
# This is my primer file D1S243 cacacaggctcacatgcc gctccagcgtcatggact D1S468 aattaaccgttttggtcct gcgacacacacttccc D1S2845 ccaaagggtgcttctc gtggcattccaacctc D1S1608 gatggcttttggggactatt cactgagccaagtgacacag D1S2893 aaaacatcaactctcccctg ctcaaaccccaataagcctt D1S2660 cacacatgcacatgcac agtgacaccagcaggg
In the example usage given above, the file 'seqs.list' contains the following sequences:
embl:Z16979 embl:Z23994 embl:Z51474 embl:G07829 embl:Z50993 embl:Z52466
Primer name D1S243 Primer name D1S468 Primer name D1S2845 Primer name D1S1608 Primer name D1S2893 Primer name D1S2660 Amplimer 1 Sequence: HSA203YC1 Z52466 H.sapiens (D1S2660) DNA segment containing (CA) repeat; clone AFMa203yc1; single read. CACACATGCACATGCAC hits forward strand at 27 with 0 mismatches AGTGACACCAGCAGGG hits reverse strand at [103] with 0 mismatches Amplimer length: 261 bp
Each primer pair consists of a name, followed by two primer sequences, primerA and primerB. The program first compares primerA to the forward strand and if it matches, primerB is compared to the reverse strand. The approach is then reversed, with the primerB being compared to the forward strand and primerA to the reverse. In this way all possible amplimers are reported.
Program name | Description |
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eprimer3 | Picks PCR primers and hybridization oligos |
stssearch | Searches a DNA database for matches with a set of STS primers |