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Prerequisites

NEXT-RNAi prerequisites depend on the mode of running that is used. E.g. for designing or evaluating siRNAs solely based on their predicted efficiency, no prerequisites are required at all. Designing or evaluating long dsRNAs requires primer3 (for primer designs) in addition. The evaluation of the reagents' specificity further requires Bowtie (for alignments). All dependencies are listed and described below.

Required software

  • For design / evaluation of long dsRNAs
  • For design / evaluation of siRNAs
    • No prerequisites

Additional software

  • Specificity evaluation of RNAi reagents (recommended) and mapping of reagents e.g. to the genome
  • Identification of low-complexity sequences (recommended)
  • Other prerequisites
    • BLAT (gapped alignments of reagents designed on CDS to the genome) (link)
    • Blast (homology evaluation of reagents) (link)
    • RNAfold (efficiency prediction using the 'rational' method) (link)

 

Installation on Debian based Linux by Installation script

The installation of NEXT-RNAi has been tested under Linux Ubuntu 9.10.

Download the installation script for Debian here. Change the script to be executable. 

cd /opt
sudo git clone https://github.com/boutroslab/nextrnai.git
sudo cp nextrnai/installation_scripts/debian/* .   
sudo chmod +x NextRnai.sh
sudo chmod +x test.sh 

Run it with administrator rights like 

sudo ./NextRNAi.sh

After installation has finished, type [y] to run the test script in order to verify the installation!

 

 

Installation on Linux manually

The installation of NEXT-RNAi has been tested under Linux Ubuntu 9.10.

All files used for this installation can be downloaded as a packed archive NEXT-RNAi_v1.4_LINUX.zip (40 MB). The packed archive includes the following files / folders:

  • nextrnai.pl (NEXT-RNAi perl script)
  • LICENCE
  • README
  • bowtie (version 0.12.5)
  • primer3 (version 1.1.1)
  • blat (suite 34)
  • blast (version 2.2.23)
  • mdust
  • ViennaRNA (version 1.8.4)

Unpack the file:

tar -xvzf NEXT-RNAi_v1.40_LINUX.tar.gz

After unpacking, please update the Next-RNAi script to the latest version from github trunk in order to always be up to date.

cd NEXT-RNAi_v1.40_LINUX;
mv nextrnai.pl nextrnai.pl.OLD 
mkdir git
git clone https://github.com/boutroslab/nextrnai.git git
cp git/NEXT-RNAi.pl nextrnai.pl  

after that the latest NEXT-RNAi and all dependencies, except for 'Vienna RNA', are ready-to-go.

The Vienna RNA package is only required by a particular efficiency calculation method ('rational' method). It can be installed the following way (requires prior installation of a gcc compiler, e.g. included in 'build-essential' package in Ubuntu):

cd software/ViennaRNA
./configure
make
sudo make install

Step-by-step instructions for downloading and installing NEXT-RNAi and dependencies for LINUX are available here.

To test NEXT-RNAi a test data-set is available for download (NEXT-RNAi_testData (236 MB)). Please download and unpack the data set in the folder where 'nextrnai.pl' is located.

tar -xvzf NEXT-RNAi_testData.tar.gz

This folder contains all files necessary to design long dsRNAs for 500 sequences from Drosophila transcripts. Options are pre-defined in 'options.txt', NEXT-RNAi just needs to be started (from within the 'testData' folder, or links in 'options.txt' need to be updated) with the command in 'start.txt'.

cd testData
perl ../nextrnai.pl -i common_regions.fa -s 250 -r d -d dmel-all-transcript-r5.26
-e NO -o options.txt -n nextrnaiTest

This will generated an 'out' folder in testData containing all results. Results can also be viewed as HTML compendium by just opening the out folder ('index.html') in any internet browser.

 

Installation on Mac OS X (10.5.X, Intel CPU)

The installation of NEXT-RNAi has been tested under MAC OS X (version 10.5.X, Intel CPU). All files used for this installation can be downloaded as a packed archive NEXT-RNAi_v1.4_MACOSX.zip (66 MB). The packed archive includes the following files / folders:

  • nextrnai.pl (NEXT-RNAi perl script)
  • LICENCE
  • README
  • bowtie (version 0.12.5)
  • primer3 (version 1.1.4-OSX)
  • blat (suite 34)
  • blast (version 2.2.23)
  • mdust (modified for running in MAC OS X)
  • ViennaRNA (version 1.8.4)

Unpack the file:

tar -xvzf NEXT-RNAi_v1.4_MACOSX.tar.gz

After unpacking, please update the Next-RNAi script to the latest version from github trunk in order to always be up to date.

cd NEXT-RNAi_v1.4_MACOSX;
mv nextrnai.pl nextrnai.pl.OLD 
mkdir git
git clone https://github.com/boutroslab/nextrnai.git git
cp git/NEXT-RNAi.pl nextrnai.pl  

after that the latest NEXT-RNAi and all dependencies, except for 'Vienna RNA', are ready-to-go.

The Vienna RNA package is only required by a particular efficiency calculation method ('rational' method). It can be installed the following way (requires prior installation of Apple Developer Tools / Xcode):

cd software/ViennaRNA
./configure
make
sudo make install

Step-by-step instructions for downloading and installing NEXT-RNAi and dependencies for MAC OS X are available here.

To test NEXT-RNAi a test data-set is available for download (NEXT-RNAi_testData (236 MB)). Please download and unpack the data set in the folder where 'nextrnai.pl' is located.

tar -xvzf NEXT-RNAi_testData.tar.gz

This folder contains all files necessary to design long dsRNAs for 500 sequences from Drosophila transcripts. Options are pre-defined in 'options.txt', NEXT-RNAi just needs to be started (from within the 'testData' folder, or links in 'options.txt' need to be updated) with the command in 'start.txt'.

cd testData
perl ../nextrnai.pl -i common_regions.fa -s 250 -r d -d dmel-all-transcript-r5.26
-e NO -o options.txt -n nextrnaiTest

This will generated an 'out' folder in testData containing all results. Results can also be viewed as HTML compendium by just opening the out folder ('index.html') in any internet browser.

Platform-independent virtual machine

The NEXT-RNAi virtual machine (running Ubuntu 9.10) allows running NEXT-RNAi via VirtualBox software that is open-source and available for all platforms (installation requires ~100MB disk space).

Ubuntu 9.10 was installed as VirtualBox virtualization. As a next step NEXT-RNAi (v1.31) and all prerequisites (Bowtie, Primer3, mdust, Blat, BLAST and ViennaRNA) were installed.

This appliance was then exported and can now be used with VirtualBox to run NEXT-RNAi (as virtualization of Ubuntu 9.10) on different platforms (including Linux, Mac, Windows).

The NEXT-RNAi virtualization can be downloaded as zipped file NEXT-RNAi_Ubuntu_VirtualBox.zip (465 MB).
The unzipped folder contains three files, a .ovf, a.mf and a .vmdk file. The .ovf file can be used to import and run the appliance with VirtualBox. User and Password for Ubuntu were both set to nextrnai. The nextrnai program 'nextrnai.pl' and all software installed are located in /home/nextrnai/NEXT-RNAi/.

To test NEXT-RNAi in the virtualization a Drosophila test data set is available for download. Click the 'xterm NEXT-RNAi' link on the Desktop. In the opened terminal use 'wget' to download the test set (236 MB):

wget http://www.dkfz.de/signaling/nextrnaiData/NEXT-RNAi_testData.tar.gz

Unpack the downloaded archive:

tar -xvzf NEXT-RNAi_testData.tar.gz

Change into the generated 'testData' folder that contains all files necessary to design long dsRNAs for 500 sequences from Drosophila transcripts. Options are pre-defined in 'options.txt'. NEXT-RNAi just needs to be started with the command in 'start.txt':

cd testData
perl ../nextrnai.pl -i common_regions.fa -s 250 -r d -d dmel-all-transcript-r5.26 -e NO
-o options.txt -n nextrnaiTest

The output is in the 'out' folder generated. By clicking on the 'NEXT-RNAi Results' link located on the Desktop a browser window will open linking to the 'testData' folder. A double-click on the out folder will open the NEXT-RNAi HTML report.

 

Detailed instructions on how NEXT-RNAi was installed with Ubuntu 9.10 in VirtualBox are available here.

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