Experimental Design
The purpose of specifying the experimental design is to provide MetaMorpheus with extra information about the spectra files you want to search, specifically each file's condition (e.g., normal or cancer tissue), the biological replicate, the fraction (if your samples are fractionated), and the technical replicate. This information is currently used to normalize intensities. In the future, we plan to extend this functionality to other aspects of MetaMorpheus (e.g., identification analysis). See the Normalization section for a description of the normalization functions.
The experimental design can be specified by clicking the "Experimental Design" button in the bottom-left part of the interface, under the mass spectra files. The experimental design window looks like this:
Requirements for each designation for experimental design are listed here:
- Condition - Not blank
- Biorep - an integer, at least 1, starting at 1. Each condition must have continuous bioreps starting at 1. For example, bioreps 1, 3, and 4 are not valid because biorep 2 is missing. In this case you would label the bioreps as 1, 2, and 3.
- Fraction - An integer, at least 1. Each biorep must have continuous fractions starting at 1. If your data is not fractionated, just enter 1 for all fractions.
- Techrep - An integer, at least 1. Each fraction must have continuous techreps starting at 1.
The experimental design window, when "Save" is clicked, saves a file called ExperimentalDesign.tsv to the folder with the first spectra file marked "Use". There is no practical reason why you would want to alter this file manually if you are using the GUI (graphical user interface). If you are using the command-line version of MetaMorpheus, however, you will need to create the .tsv file yourself manually. An example file is attached on the bottom of this page. Be sure to follow the "Formatting" rules.
Currently, the experimental design settings are only used for intensity (i.e., quantification) normalization. In the future, we plan to use these experimental design settings for PSM analysis.