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Viral phylogenetics: Move -gtr, -gamma params back up
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src/content/viral-phylogenetics/steps/Step3.md

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@@ -8,26 +8,21 @@ Now that we have run MSA on our dataset, we can perform Phylogenetic Inference:
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We will use [FastTree](https://morgannprice.github.io/fasttree/) to generate an unrooted phylogenetic tree to assess relative ancestral relationships between our samples.
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1. Try <Execute command="FastTree -help" inline /> to take a look at the usage instructions.
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1. Try <Execute command="FastTree" inline /> to take a look at the usage instructions.
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2. Now, generate our phylogenetic tree:
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<Execute command="FastTree -nt ViralMSA_Out/sarscov2_sequences.fas.aln > sarscov2_sequences.unrooted_tree.nwk" />
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<Execute command={"FastTree \\ -nt \\ -gtr \\ -gamma \\ ViralMSA_Out/sarscov2_sequences.fas.aln > sarscov2_sequences.unrooted_tree.nwk"} />
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Let's make some sense of this command:
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- `-nt` specifies that our alignment is of **n**ucleo**t**ides and not amino acids
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- `sarscov2_sequences.msa.fas > sarscov2_sequences.unrooted_tree.nwk` tells FastTree to take in our multiple sequence alignment file (from Step 1) as input and to output the unrooted phylogenetic tree to a file called `sarscov2_sequences.unrooted_tree.nwk` in the same directory. A `.nwk` file is in Newick format, which is often used to represent phylogenetic trees. It is a text-based way to represent the tree structure. You can read more about Newick format [here](https://en.wikipedia.org/wiki/Newick_format).
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**Optional Options:**
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Note: The following options may be included in the command to improve accuracy at the cost of increased runtime.
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- `-gtr` implements use of a [Generalized Time-Reversible](https://en.wikipedia.org/wiki/Substitution_model#Generalised_time_reversible) (GTR) model of evolution for our tree. FastTree can be run with either the [Jukes-Cantor](https://en.wikipedia.org/wiki/Models_of_DNA_evolution#JC69_model_(Jukes_and_Cantor_1969)) or GTR model.
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- `-gamma` allows for rescaling of the branch lengths and computation of a Gamma2-based likelihood
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- `sarscov2_sequences.msa.fas > sarscov2_sequences.unrooted_tree.nwk` tells FastTree to take in our multiple sequence alignment file (from Step 1) as input and to output the unrooted phylogenetic tree to a file called `sarscov2_sequences.unrooted_tree.nwk` in the same directory. A `.nwk` file is in Newick format, which is often used to represent phylogenetic trees. It is a text-based way to represent the tree structure. You can read more about Newick format [here](https://en.wikipedia.org/wiki/Newick_format).
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3. Use `head` to view the first 10 lines of the tree file:
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