====== Samtools, Tophat and Tophat2 ======

A spliced read mapper for RNA-Seq.
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  * **SAMTOOLS**
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Download
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<code>
# wget https://sourceforge.net/projects/samtools/files/latest/download?source=files -O samtools-1.3.1.tar.bz2
# tar xvjf samtools-1.3.1.tar.bz2
</code>
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    * Compile and Install
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<code>
# cd samtools-1.3.1/
# ./configure
# make
# make install
</code>
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    * Check
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<code>
$ samtools --version

samtools 1.3.1
Using htslib 1.3.1
Copyright (C) 2016 Genome Research Ltd.
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  * **TOPHAT**
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Download

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<code>
# wget http://ccb.jhu.edu/software/tophat/downloads/tophat-1.4.1.Linux_x86_64.tar.gz
# tar xvzf tophat-1.4.1.Linux_x86_64.tar.gz
</code>
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    * Install
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<code>
# mv tophat-1.4.1.Linux_x86_64 /usr/local/bin/
# ln -s /usr/local/bin/tophat-1.4.1.Linux_x86_64/tophat /usr/local/bin/
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    * Test
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<code>
$ wget http://ccb.jhu.edu/software/tophat/downloads/test_data.tar.gz
$ tar zxvf test_data.tar.gz
$ cd test_data/
$ tophat -r 20 test_ref reads_1.fq reads_2.fq


[Thu Jan 12 19:49:46 2017] Beginning TopHat run (v1.4.1)
-----------------------------------------------
[Thu Jan 12 19:49:46 2017] Preparing output location ./tophat_out/
[Thu Jan 12 19:49:46 2017] Checking for Bowtie index files
[Thu Jan 12 19:49:46 2017] Checking for reference FASTA file
[Thu Jan 12 19:49:46 2017] Checking for Bowtie
        Bowtie version:                  1.1.2.0
[Thu Jan 12 19:49:46 2017] Checking for Samtools
Error: TopHat requires Samtools 0.1.7 or later
</code>
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**NOTE**
samtools 1.3.1 so samtools check error

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  * **TOPHAT2**
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Download

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<code>
# wget http://ccb.jhu.edu/software/tophat/downloads/tophat-2.1.1.Linux_x86_64.tar.gz
# tar xvfz tophat-2.1.1.Linux_x86_64.tar.gz 
</code>
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  * Install
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<code>
# mv tophat-2.1.1.Linux_x86_64 /usr/local/bin/ 
# ln -s /usr/local/bin/tophat-2.1.1.Linux_x86_64/tophat2 /usr/local/bin/
# chown -R root:root /usr/local/bin/tophat-2.1.1.Linux_x86_64  
</code>
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  * Test
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<code>
$ wget http://tophat.cbcb.umd.edu/downloads/test_data.tar.gz 
$ tar zxvf test_data.tar.gz 
$ cd test_data/
$ tophat2 -r 20 test_ref reads_1.fq reads_2.fq

[2017-01-12 19:53:31] Beginning TopHat run (v2.1.1)
-----------------------------------------------
[2017-01-12 19:53:31] Checking for Bowtie
                  Bowtie version:        2.2.9.0
[2017-01-12 19:53:31] Checking for Bowtie index files (genome)..
        Found both Bowtie1 and Bowtie2 indexes.
[2017-01-12 19:53:31] Checking for reference FASTA file
[2017-01-12 19:53:31] Generating SAM header for test_ref
[2017-01-12 19:53:31] Preparing reads
         left reads: min. length=75, max. length=75, 100 kept reads (0 discarded)
        right reads: min. length=75, max. length=75, 100 kept reads (0 discarded)
[2017-01-12 19:53:31] Mapping left_kept_reads to genome test_ref with Bowtie2
[2017-01-12 19:53:32] Mapping left_kept_reads_seg1 to genome test_ref with Bowtie2 (1/3)
[2017-01-12 19:53:32] Mapping left_kept_reads_seg2 to genome test_ref with Bowtie2 (2/3)
[2017-01-12 19:53:32] Mapping left_kept_reads_seg3 to genome test_ref with Bowtie2 (3/3)
[2017-01-12 19:53:32] Mapping right_kept_reads to genome test_ref with Bowtie2
[2017-01-12 19:53:32] Mapping right_kept_reads_seg1 to genome test_ref with Bowtie2 (1/3)
[2017-01-12 19:53:32] Mapping right_kept_reads_seg2 to genome test_ref with Bowtie2 (2/3)
[2017-01-12 19:53:32] Mapping right_kept_reads_seg3 to genome test_ref with Bowtie2 (3/3)
[2017-01-12 19:53:32] Searching for junctions via segment mapping
[2017-01-12 19:53:32] Retrieving sequences for splices
[2017-01-12 19:53:32] Indexing splices
Building a SMALL index
[2017-01-12 19:53:33] Mapping left_kept_reads_seg1 to genome segment_juncs with Bowtie2 (1/3)
[2017-01-12 19:53:33] Mapping left_kept_reads_seg2 to genome segment_juncs with Bowtie2 (2/3)
[2017-01-12 19:53:33] Mapping left_kept_reads_seg3 to genome segment_juncs with Bowtie2 (3/3)
[2017-01-12 19:53:33] Joining segment hits
[2017-01-12 19:53:33] Mapping right_kept_reads_seg1 to genome segment_juncs with Bowtie2 (1/3)
[2017-01-12 19:53:33] Mapping right_kept_reads_seg2 to genome segment_juncs with Bowtie2 (2/3)
[2017-01-12 19:53:33] Mapping right_kept_reads_seg3 to genome segment_juncs with Bowtie2 (3/3)
[2017-01-12 19:53:34] Joining segment hits
[2017-01-12 19:53:34] Reporting output tracks
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[2017-01-12 19:53:34] A summary of the alignment counts can be found in ./tophat_out/align_summary.txt
[2017-01-12 19:53:34] Run complete: 00:00:02 elapsed
</code>
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**NOTE**
Tophat2 v2.1.1 test_data error with bowtie2 v2.3.0