primerfinder

This program integrates primer3 and OligoCalc functions to pick candidate primers for staining.

Workflow

Take RNA sequence as input
Call Primer3 to find hybridization probe (internal oligo) candidates with 21 bases with melting temperature 45~65 (celcius)
Pick candidates ending with “TA” or “AG”
Generate DNA primer pairs (20+20) for valid candidates
Use the OligoCalc algorithm to check the melting temperature and self-complementarity of primer pairs
Output valid primer pairs

Download the entire "primerfinder" project to local machine. Creat a folder named "primer3" under the project folder. Unzip the "primer3.zip" file and place all primer3 files directly under the primer3 folder.
Open Windows command line
Go to the project folder
Save nucleotide sequence in a file in the input folder, such as “.\input\SCGB1A1.txt”
Make sure Rscript.exe is in your system path
Run the program by typing “Rscript primerfinder.R .\input\SCGB1A1.txt”
The output will be saved in a “uidxxx” folder in output directory

File	Description
primer3.in.txt	the generated input file for primer3
primers.1.all.txt	all primer candidates returned by primer3
primers.2.validending.txt	all primer candidates with valid ending
primers.3.pairs.txt	all primer pairs
primers.4.final.txt	valid primer pairs

Currently support Windows system.
Currently only accept bases: A, C, T, G, U.
The program is in the testing phase. Highly recommend double checking the properties of the final primers in OligoCalc (http://biotools.nubic.northwestern.edu/OligoCalc.html).
The implementation of the self-complementarity checking algorithm is more stringent than the implementation in OligoCalc.
The Windows executables of primer3 (libprimer3 release 2.3.6) were downloaded from http://primer3.sourceforge.net/.
The primer3 execution results are consistent with this web version of primer3: http://primer3.ut.ee/.