#Metagenome binning using MetaBAT Authored by Fan Yang EDAMAME-2016 wiki
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##Overarching Goal
- This tutorial will contribute towards an understanding of microbial metagenome analysis
##Learning Objectives
- Understand metagenomic binning and perform a binning exercise
##Tutorial We will be running a specific tool for metagenomic binning called MetaBAT. You can read about this tool and its applications in the original paper, [MetaBAT, an efficient tool for accurately reconstructing single genomes from complex microbial communities] (https://peerj.com/articles/1165/)
Let's start by getting our computer!
- Initiate an Amazon EC2 instance if you don't have one launched already
a. ubuntu
b. m3.large
Now, let's download our software.
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Where to get MetaBAT?
A: MetaBAT is deposited in BitBucket (remote repository like GitHub)cd ~ sudo apt-get install git ## EC2 does not have git installed by default
git clone https://bitbucket.org/berkeleylab/metabat.git metabat_clone git clone https://github.com/edamame-course/metagenome_binning
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How to install MetaBAT? Do I have all softwares I need to run MetaBAT?
cd metabat_clone less README.md less INSTALL.md
Don't forget to push "q" key to exist the "less" window.
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Install MetaBAT
cd ~ bash ~/metagenome_binning/installation.sh
This tutorial takes place assuming you have an assembled metagenome and its mapped read abundances.
- So you learned about how to assemble sequences yesterday and assembled some contigs,
for today's tutorial, we need a full assembled file.
Let's also get some assembled and mapped data.
cd ~ mkdir data && cd ~/data wget https://s3.amazonaws.com/edamame/SRR492066.sam.bam.sorted.bam wget https://s3.amazonaws.com/edamame/SRR492065.sam.bam.sorted.bam wget https://s3.amazonaws.com/edamame/final.contigs.fa
Q: Can you think of a reason why we couldn't use the partial assembled contigs from the metagenome assembly exercise?
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Now let's do some binning!
cd ~/data for i in *.sorted.bam do mkdir ~/data/$i.bins cd ~/data/$i.bins ~/metabat/bin/runMetaBat.sh ~/data/final.contigs.fa ~/data/$i done
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What if we want to bin the contigs with different threshold?
cd ~/data/SRR492065.sam.bam.sorted.bam.bins #First, try sensitive mode to better sensitivity ~/metabat/bin/metabat -i ~/data/final.contigs.fa -a final.contigs.fa.depth.txt -o bin1 --sensitive -l -v --saveTNF saved.tnf --saveDistance saved.gprob #Try specific mode to improve specificity further; this time the binning will be much faster since it reuses saved calculations ~/metabat/bin/metabat -i ~/data/final.contigs.fa -a final.contigs.fa.depth.txt -o bin2 --specific -l -v --saveTNF saved.tnf --saveDistance saved.gprob #Try specific mode with paired data to improve sensitivity while minimizing the loss of specificity ~/metabat/bin/metabat -i ~/data/final.contigs.fa -p final.contigs.fa.paired.txt -o bin3 --specific -l -v --saveTNF saved.tnf --saveDistance saved.gprob