helps to plan a RFLP experiment: It finds restriction enzymes
that cut a set of homologue DNA sequences in different ways and simulates the resulting gel- electrophoresis image. The program helps you to find the crucial restriction enzymes that characterize the population. With the electrophoresis simulation it is easy to chose the restriction enzymes that show the best polymorphism on the gel.
An important keyword is biodiversity
. The program is written in perl
, uses some modules of bioperl
and the GD-Library
. The program should be platform independet, but is tested only on Mac OS X.
Different homologue sequences, representative for the investigated population. For example, some sequences of PCR products, obtained with the same primer pair that you wish to use in your RFLP experiment (sample file
). The sequences format should be accepted from bioperl modules (Fasta, EMBL, GenBank, raw, .. ). All sequences should be in the same file.
- Simulation of gelimages (PNG): Every sequence one image. Every image with a molecular weight marker. Every restriction enzyme one slot.
- Evaluation is done with the whole bioperl restriction enzyme database, but desired enzymes also can be added (EnzymeNAME or RESTRICTION^SITE)
- Isoshizomers are not used for calculations (but appear in the resulting fragments.txt)
- Sequences can be trimmed to the same lenght or not, but for the gelimage the original sequences are used
- Desired molecular weight marker bands can be labelled
- Option to label images (with sequence name)
- Lots of different in- and output formats are supported (bioperl)
- Abuse the program to cut primer sequences of a bunch of sequences:
- 1. in one sequence cut primer sequence manually
- 2. import all sequences (fasta)
- 3. choose: trimm = 1, aligned_format = 'fasta'
- Automatic remove bad aligning sequences
- Chose range of searched restriction enzymes (4, 6, .. -bp cutter)
- Automatic reverse complement sequences (does not work for large amount of seq; if it does not work try with fewer sequences, exoprt alignment in fasta, join all sequences again, calculate the whole)
- Animate sequence of images (X-Windows and ImageMagick)
$> animate -delay 25 imagepath/*.png
- Make an anim gif (ImageMagick), as in example above
$> convert -delay 40 path/to/images/*.png anim.gif
Author and Mehladress
- Add option to mark primers on gel (red / blue dyes)
- Add option to remove primers, primers could be added with an extra file (fw------back)
- Beautify the code: Make some more variables local
- Faster calculations: Do calculations only once for identical sequences
Alignement and restriction
- Term::ANSIColor #only for output a red nucleotide in cli, obsolete:)
- man Bio::Tools::RestrictionEnzyme
- man Bio::Align::AlignI
- man Bio::AlignIO
- man Bio::Seq
- man Bio::SeqIO
- man Bio::Tools::Run::Alignment::Clustalw