G-POT Primer Set Selection Tool, rev. Nov. 8, 2006
====================================================

Description
-----------

G-POT is a primer set selection tool that minimizes the number of
primers needed to amplify a set of loci while ensuring that each
locus is amplified by a pair of primers located within L basepairs
of each other, where L is an input parameter.  The tool is based
the "potential function" greedy algorithm described in:

K.M. Konwar, I.I. Mandoiu, A.C. Russell, and A.A. Shvartsman.  
Improved Algorithms for Multiplex PCR Primer Set Selection with
Amplification Length Constraints, Proc. 3rd Asia-Pacific
Bioinformatics Conference (APBC), pp. 41-50, 2005.


Building G-POT
-------------

The G-POT package contains the source code for two executables:
 * g-pot, the primer selection algorithm
 * gen, a random instance generator used for testing

To build the two executables execute the following commands in the directory 
where g-pot.1.0.tar.gz is located:

tar -xzvf g-pot.1.0.tar.gz
cd g-pot.1.0
make all

After compiling the executables, make runs a regression test (assuming that awk 
is installed on your system) to verify that they work as expected.  Upon 
completion of this test you should get the message "REGRESSION TEST 
SUCCESSFUL!!!"

The code has been compiled and tested successfully on Gentoo
Linux with gcc compiler version 3.3.5 and Red Hat 9 with gcc
compiler version 3.3.1.


Using G-POT
-----------

G-POT is a command-line tool which by default reads from the standard input 
and prints the results to the standard output.

Usage:
 g-pot [options] [< input_file] [> output_file]


OPTIONS
-plen N           : primer length (default: 10)
-prox N           : minimum distance between primer and amplification locus (default: 0)
-prodlen N        : maximum product length (default: 1000)
-max_melt_temp N  : maximum primer temperature (default: none)
-min_melt_temp N  : maximum primer temperature (default: none)
-min_gc N         : minimum gc content of the primers (default: 0 %)
-max_gc N         : maximum gc content of the primers (default: 100 %)
-salt_conc N      : salt concentration in the reaction (default: 50 milimolar)
-dna_conc N       : dna concentration in the reaction (default: 50 nanomolar)


Input Format
------------

The input file format for each line is <sequence1,sequence2> where
sequence1 and sequence2 are the 5'-3' sequences flanking the
corresponding amplification target. See the included
"sample_input" file for a sample.

Output Format
-------------

By default G-POT prints the results to the standard output.  The
output includes the parameters used for the program, the runtime, 
the number of selected primers, the number of covered targets, 
and the list of 5'-3' primer sequences including primer melting 
temperature and GC content.  The output of the following command:

./g-pot -plen 17 -min_melt_temp 55 -max_melt_temp 60 -input sample_input

can be found in file "sample_output".


Using GEN
-----------

The random sequence generator can be used as follows:
./gen [-m M] [-n N] [-seed S]

where: 
 * M is the number of amplification targets (default 10)
 * N is the length of the flanking sequences (default 10) 
 * S is the random number generator seed (default 1)

For example:

./gen -n 10 -m 5 -seed 4

would output

< cgctacttta , agccggccgc >
< tcgaggggac , cagggcagct >
< agcaattcga , tggggacgat >
< aagttgttaa , gctctgtaac >
< ttttctgatc , cgcgaagtgg >