5. Tree building

Introduction

We can now use our supermatrix to build a phylogeny. Here we will be using FastTree to quickly generate a tree, since this is (unsurprisingly) fast. This is an “approximately maximum likelihood” phylogenetic tree building method, broadly similar in method to PhyML, RAxML or IQtree. It is generally a little less accurate, but it’s faster so better for fiddling around with and learning on.

Data and software

The input data for this tutorial is a supermatrix of a set of sequences for which you want to build a phylogeny, and a table of taxonomy for those sequences. If you’re following along step-by-step, the supermatrix was produced in the previous tutorial. The file taxonomy_GB_CCCP.csv within the sectionD archive is the corresponding taxonomy table.

This tutorial uses the FastTree software, as well as the phylabel.R script. You should also have a tree viewer, such as FigTree or Dendroscope, installed on your personal computer.

Running FastTree

Use the following command, using the supermatrix output from the previous step as the input and choosing an appropriate name for your tree. The -gtr option tells FastTree to use the GTR+CAT model, and the -nt option specifies that this is DNA data.

FastTree -gtr -nt < input.fasta > output.tre

The algorithm generates a newick file, which is the most common format for phylogenetic trees. This is a plain text format, you can view the file using the head command. Each terminal in the tree has the ID of the sequence used to generate it.

Adding taxonomy

The terminals of the tree are named only with the code names for their respective source sequences. We would like to add in the taxonomy so that the terminals are meaningful. There are lots of ways to do this, but our usual method is to use a custom script which takes as input the tree and a table that records the taxonomy of all of the sequences. Happily enough, we happen to have that table handy. Run the following command to rename the tree tips with taxonomy:

phylabel.R -p input.tre -r -y taxonomy.csv -o output.tre

Assessing Trees

You can download the renamed tree to your computer and open it using a tree viewer such as Figtree or Dendroscope. If you’re completely new to viewing phylogenetic trees, you may want to read the page on viewing trees.

Exercise

First we should make sure our tree is correctly rooted. Our most basal clade is some sequences of Araneae, so these three sequences should be sisters to the rest of the tree. Find the clade containing these three terminals and set it as the outgroup (see here if you’re not sure how to reroot the tree). Make sure to reroot both your renamed and un-renamed versions.

Whenever we build a phylogenetic tree, we must closely inspect it to determine how well the tree building performed. Have a look at your tree for features that aren’t quite right:

• Check to see whether, in general, the taxonomy of your terminals matches the clade structure of the tree
• Check to see whether there are any very long branches. Why might these occur?

Next Steps

If you’ve been following through these steps with your own data, we would suggest also trying out a more thorough tree building program such as RAxML, IQtree, PhyML, MrBayes or BEAST. These are much slower to run with mitogenome-scale data, but are much more accurate and appropriate for building a tree for a real analysis. Once you have your tree, you can go back to the tutorial on Phylogenetic placement to place your OTUs within your tree.