SAT� is a method for estimating multiple sequence alignments and trees that has been shown to produce highly accurate results for datasets with large numbers of sequences. Running SAT� using its default settings is very simple, but improved accuracy can be obtained by modifying its algorithmic parameters. We provide a detailed introduction to the algorithmic approach used by SAT�, and instructions for running a SAT� analysis using the GUI under default settings. We also provide a discussion of how to modify these settings to obtain improved results, and how to use SAT� in a phylogenetic analysis pipeline.