Scripts to aid the design of differential primers for diagnostic PCR.
Python
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README
find_differential_primers.py

README

This repository contains code for finding discriminatory primers among genomes 
of interest. Please refer to the following:

CITATION (method and application to human bacterial pathogens, sub-serotype resolution):
Pritchard L, Holden NJ, Bielaszewska M, Karch H, Toth IK (2012) "Alignment-Free 
Design of Highly Discriminatory Diagnostic Primer Sets for Escherichia coli 
O104:H4 Outbreak Strains." PLoS ONE 7(4): e34498. doi:10.1371/journal.pone.0034498

CITATION LINK:
http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0034498

or 

CITATION (application to plant pathogens, species-level resolution)
Pritchard L, Humphris S, Saddler GS, Parkinson NM, Bertrand V, Elphinstone JG, 
Toth IK (2013) "Detection of phytopathogens of the genus Dickeya using a PCR primer 
prediction pipeline for draft bacterial genome sequences." Plant Pathology, 62, 587-596
doi:10.1111/j.1365-3059.2012.02678.x

CITATION LINK
http://onlinelibrary.wiley.com/doi/10.1111/j.1365-3059.2012.02678.x/full

DEPENDENCIES:
Any of the following Dependencies can be used for running the 
'find_differential_primers.py' pipeline:

>> biopython : v1.57 - v1.61
Git Hub version : https://github.com/biopython/biopython
Older Releases (scroll to bottom): http://biopython.org/wiki/Download

>> bx-python : current Hg (Mercurial) version works as of June 19, 2013
Hg version : https://bitbucket.org/james_taylor/bx-python/src

>> EMBOSS (for eprimer3) : v6.30, v6.31, v6.4.0
http://emboss.sourceforge.net/download/

>> primer3  : v1.1.4
http://primer3.sourceforge.net/releases.php

>> prodigal  : v1.20
http://code.google.com/p/prodigal/downloads/list?can=1&q=&colspec=Filename+Summary+Uploaded+ReleaseDate+Size+DownloadCount

>> BLAST / BLAST+ : v2.2.22 / v2.2.22+  (Other versions may work)
http://blast.ncbi.nlm.nih.gov/Blast.cgi?CMD=Web&PAGE_TYPE=BlastDocs&DOC_TYPE=Download


BASIC USE:
If you do not have complete genome sequences use the 'stitch_six_frame_stops.py'
script to stitch together the contigs of a single genome (contained within one
multi-FASTA formatted file). The script does this by stitching the contigs with
the following sequence 'NNNNNCATTCCATTCATTAATTAATTAATGAATGAATGNNNNN'. This
sequence contains start and stop codons in all frames. Remember the default 
workflow is to only find primers within CDS regions.

Once all genomes have been stitched together, make a config file similar
to the example within the 'O104_primers_5.conf' file.

Finally, run 'find_differential_primers.py'.

FURTHER INFORMATION:
Read the comments contained within the top of each '*.py' file as well as the 
Supporting Information ('Methods S1' document) of the above CITATION.

CONTRIBUTORS
============
Leighton Pritchard   https://github.com/widdowquinn
Benjamin Leopold     https://github.com/cometsong
Michael Robeson      https://github.com/mikerobeson