You can simply obtain this entire workflow by cloning the Code4Rice3K repository into your local machine as follows:
git clone https://github.com/BrendelGroup/Code4Rice3K
cd Code4Rice3KThis workflow has been developed and tested on Linux machines only.
Specifically, Ubuntu and Red Hat Enterprise linux systems.
That being said, there are other softwares and tools that need to be installed on your machine (if they are not already installed) as they are intrinsic to this workflow.
For convenience, a script is provided under the bin/ directory to check for any required software.
You can simply run it with ./xinstallcheckprerequisites, and it should tell you if if finds an error.
There is more than one way to install softwares on your linux machine;
You can run the xinstallCode4Rice3K script provided under the bin/ directory, it will install all the necessary softwares.
Or, if you prefer, you can manually install these softwares using the short guide below.
For either methods, please make sure to keep track of paths of any installed binaries (see below for details).
The versions listed are the ones that have been successfully tested.
See https://www.java.com/en/download/ for details. Last update: August 1, 2017.
You can check the availability and version of java on your machine by running:
java -version
If you need to install it, please follow the instructions below:
mkdir $YOUR_PREFERRED_DIRECTORY/Java
cd Java
wget -c --header "Cookie: oraclelicense=accept-securebackup-cookie" http://download.oracle.com/otn-pub/java/jdk/8u131-b11/d54c1d3a095b4ff2b6607d096fa80163/jdk-8u131-linux-x64.tar.gz
tar zxvf jdk-8u131-linux-x64.tar.gz
# Now use an editor like "vim" or "nano" to add the following lines into your "~/.profile", save it, and run "source ~/.profile" in the command line:
export JAVA_HOME=path_to_jdk1.8.0_131_directory
export PATH=$JAVA_HOME/bin:$PATHSee http://www.htslib.org/download/ for details. Last update: August 1, 2017.
First, check the availability and version of samtools on your machine. You can do this by running:
samtools --help
If you want to install it, use:
mkdir $YOUR_PREFERRED_DIRECTORY/Samtools
cd Samtools
wget https://github.com/samtools/samtools/releases/download/1.5/samtools-1.5.tar.bz2
tar xvjf samtools-1.5.tar.bz2
cd samtools-1.5/
./configure --without-curses --disable-lzma
make
make install # You may need to use "sudo make install"See http://www.htslib.org/doc/tabix.html for details. Last update: August 1, 2017.
First, check the availability and version of tabix on your machine. You can do this by:
tabix --help
If you want to install it, use:
mkdir $YOUR_PREFERRED_DIRECTORY/Tabix
cd Tabix
wget https://sourceforge.net/projects/samtools/files/tabix/tabix-0.2.6.tar.bz2
tar xvjf tabix-0.2.6.tar.bz2
cd tabix-0.2.6
make
# add the following line to your "~/.profile" file found in your home directory, save the file, and then run "source ~/.profile" in the command line.
export PATH=$PATH:/path_to_tabix_dir/tabix-0.2.6See http://www.htslib.org/download/ for details. Last update: August 1, 2017.
First, check the availability and version of bcftools on your machine. You can do this by running:
bcftools --help
If you want to install it, use:
mkdir $YOUR_PREFERRED_DIRECTORY/Bcftools
cd Bcftools
wget https://github.com/samtools/bcftools/releases/download/1.5/bcftools-1.5.tar.bz2
tar xvjf bcftools-1.5.tar.bz2
cd bcftools-1.5/
./configure --without-curses --disable-lzma
make
make install # You may need to use "sudo make install"See http://vcftools.sourceforge.net for details. Last update: August 1, 2017.
First, check the availability and version of vcftools on your machine. You can do this by running:
vcftools --help
If you want to install it, first you have to install Tabix (see above) then:
mkdir YOUR_PREFERRED_DIRECTORY/VCFtools
cd VCFtools
wget https://sourceforge.net/projects/vcftools/files/vcftools_0.1.13.tar.gz
tar zvxf vcftools_0.1.13.tar.gz
cd vcftools_0.1.13
make
# add the following line to your "~/.profile" file found in your home directory, save the file, and then run "source ~/.profile" in the command line.
export PERL5LIB=/path_to_vcftools/vcftools_0.1.13/perl
export PATH=$PATH:/path_to_vcftools/vcftools_0.1.13/bin/See https://www.python.org for details. Last update: August 1, 2017.
You can check the availability and version of bcftools on your machine by running:
python --version
Python usually come pre-installed on most machines. If that is not the case and you want to install it, then there is more than one way to do so. Check the Python.org website for installation instructions.
See http://biopython.org for details. Last update: August 1, 2017.
If you have administrative privilage access on your machine, use the package manager for best results. Otherwise, we recommend using pip.
pip install biopython --userSee https://pypi.python.org/pypi/PyVCF for details. Last update: August 1, 2017.
Same as installing biopython, you can use:
pip install pyvcf --userSee https://broadinstitute.github.io/picard/ for details. Last update: September 15, 2017.
mkdir $YOUR_PREFERRED_DIRECTORY/Picard
cd Picard/
wget https://github.com/broadinstitute/picard/releases/download/2.10.9/picard.jarYou will need to set the path for Picard in the xsetupReference script under the bin/ directory.
Open the script using an editor, and change the path in the following line to where Picard is installed:
PICARD=/usr/local/src/NGS_DIR/Picard/picard.jarSee https://software.broadinstitute.org/gatk/ for details. Last update: September 15, 2017.
mkdir $YOUR_PREFERRED_DIRECTORY/GATK
cd GATK
wget -qO- 'https://software.broadinstitute.org/gatk/download/auth?package=GATK-archive&version=3.6-0-g89b7209' | tar xfj -You will need to set the path for GATK in the code4Rice3K.conf configuration file under the bin/ directory.
Open the file using an editor, and change the path in the following line to where GATK is installed:
gatk=java -jar /usr/local/src/code4Rice3K/GATK/GenomeAnalysisTK.jarSee RAxML for details. Last update: August 1, 2017.
First, check the availability and version of RAxML on your machine. You can do this by running:
raxmlHPC-PTHREADS -help
For installations, you will have to clone the RAxML repository from github:
mkdir $YOUR_PREFERRED_DIRECTORY/RAxML
cd RAxML
git clone https://github.com/stamatak/standard-RAxML.git
cd standard-RAxML/
make -f Makefile.gcc
rm *.o
# add the following line to your "~/.profile" file found in your home directory, save the file, and then run "source ~/.profile" in the command line.
export PATH=$PATH:/path_to_raxml_directory/raxmlHPC-PTHREADSThe only step left to be done is to prepare the reference genome for the variant calling.
You can do this by executing the xsetupReference in the bin/ directory, which will install the reference/ directory in your code4Rice3K install directory.