MysiRNA source code for Academic Users, Version 1 ===================================================== ------------- License Terms ------------- MysiRNA-Designer a Software for Rational siRNA Design Copyright (C) 2011 This program is free software; you can redistribute it and/or modify it under the terms of the GNU General Public License as published by the Free Software Foundation; either version 2 of the License, or (at your option) any later version. This program is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for more details. You should have received a copy of the GNU General Public License along with this program; if not, write to the Free Software Foundation, Inc., 59 Temple Place, Suite 330, Boston, MA 02111-1307 USA see the Copyrights files inclosed for this software and other excutables COPYING. ------------- Requirements ------------- Windows envirenment -PPM (Perl Package Manager) varsion (5.10.1.1008): avaible at (http://www.activestate.com/activeperl/downloads), to be installed. -Bioperl module follow the "GUI Installation" instructions stated for version 5.10 at (http://www.bioperl.org/wiki/Installing_Bioperl_on_Windows). After finishing the installation of "Bioperl" install these additional packages: "bioperl-Run", "Bundle-bioPerl","Modern-Perl" and "spreadsheet-ParseExcel" by typing their named in the search box of PPM, mark them for install and install them using the arrow-shaped install icon. -EMBOSSwin: available at (http://www.interactive-biosoftware.com/embosswin/embosswin.html). after installation define a system varriable named "EMBOSSWIN" by accessing windows "advanced system setting"->"Environment Varriable". Under "system varriables" add new varriable named "EMBOSSWIN" and give it the PATH where EMBOSSwin is installed. -Download mRNA dataset (refseq) from NCBI website (for off-target filtration, optional): (ftp://ftp.ncbi.nih.gov/blast/db/) named "refseq_rna.tar.gz" extract it using decompressing tools as "winRAR" or "7_Zip". -WEKA 3.7: Download and install the software keeping the Path of the installation known. WEKA is a freeware available online at (http://www.cs.waikato.ac.nz/ml/weka/) -------------------------------- Before Running MysiRNA Designer -------------------------------- -Enable Excel macros by changing the macro security level, Access "Excel Option"-> "Trust Centre"->"Trust Centre Setting"->"Macro settings"->"Enabl all macros". -Check EMBOSSWIN environment variable whether it has been defined or not via accessing "Advanced System Setting"->"Environment Varriables"->add in the System Varriables. Search for EMBOSSWIN system varriable, if it does not exist add them as per below: Varriable name: EMBOSSWIN Varriable Value: (emboss installation path, detault: "C:\embosswin"). -Access Perl Package Maneger "PPM" installed packages and check if following package are successfully installed: (BioPerl, bioperl-Run, Bundle-bioPerl, Modern-Perl and spreadsheet-ParseExcel). ------------------------ Running MysiRNA Designer ------------------------ Double click MysiRNA_Designer.exe Syntax: (perl) MysiRNA_Designer.pl -s -o -w Use the Following Mandatory Options:\n -s File containing mRNA(s) Refseq Accession(s) one/line, -o out put PATH. -w WEKA installation PATH. Use the Following Non-Mandatory Options:\n -d To perform off-target filtration, insert mRNA refseq dataset name without its extension and complete PATH, . -t RNAxs threshold on the 8 nts accessibility, else use default. -u RNAxs threshold on the 16 nts accessibility, else use default. -p Use MysiRNA-Model high specificity threshold (Y/N). -e Updated 3UTR Database, else use default -i Updated refseq2ensumble ID Database, else use default For Queries about the installaion, type \'notepad README.txt\' For Queries about the Copy rights, type \'notepad COPYING.txt\' - To see the program help type MysiRNA_Designer.pl (with no parameters). the program will be producing the resutls in a folder name results and the final results in a folder named total_mysirna - The program can accept multiple mRNA(s) accession at single run. ------------ Output Files ------------ The MysiRNA program generates numerous text output files in varrious folders: ------------------------------------------ "MysiRNA_Designer_Final_results" contains: ------------------------------------------ - X_Final_siRNA.fasta (Final siRNA passing MysiRNA_Designer filtration phases, X indicate mRNA order in the intry file). - X_Final_MysiRNA.fasta (same siRNA as X_Final_siRNA.fasta after being evaluated with MysiRNA model, X indicate mRNA order in the intry file). ------------------- "tools_results" contains: ------------------- - fasta1results.fasta (mRNA Sequence in fasta format of Accession entered), - fasta1result.gb (Genbank format of the accession entered), - blast1result.fasta (the accession of all the transcripts of the mRNA), - Tar_acc_result.txt (target accessible siRNA), - SNPs_free_siRNAs.fasta (siRNA free from SNPs), - snpsFinal.fasta (SNPs regions), --------------------------------------------- "blastresult" contains off-target evaluation: --------------------------------------------- - siRNA_num_X_Off_Target.fasta (Blast results siRNAs where X change by siRNA number) - X_Whole_siRNA_off_targets.csv (all the blast results for all siRNA, where X change by mRNA number). -------------------------------- "Multi_score_Accepted" contains: -------------------------------- - X_Multi_score_Approved_siRNA.fasta (siRNA passing multiple scores, multiple transcripts and SNPs filtration). ---------------------------------------------------------------------------------- general messages from standard output/error to follow the program stages/errors. ------- Testing ------- Type: Perl mysiRNA_Designer.pl -o -w -s Where, acc.txt use a file containing the accession: NM_001667 After complete run, the result should appear as it is presented in Test_Results.txt With 22 possible siRNA targeting NM_001667 mRNA. Developed by M.Mysara et al. Mysara M, Garibaldi J, ElHefnawi M,(2011) “MysiRNA-Designer: A Workflow For Efficient siRNA Design”, PloS One Journal. if you have any question contact: mm.abdelwahab@nrc.sci.eg, mohamed.mysara@nottingham.ac.uk and mohamed.abdelwahab@mail.iti.net.eg