Pulsar is the search engine developed at Biognosys. It is integrated into Spectronaut™ to make the workflow as smooth and seamless as possible. With Pulsar, complete data independent acquisition (DIA) workflows can be performed within one software. Pulsar has two main functionalities: 1) generating spectral libraries and 2) performing directDIA™.
1. Generating libraries
Notably, Pulsar is not only able to search data dependent acquisition (DDA) runs but also DIA and parallel reaction monitoring (PRM) data, allowing the generation of libraries from most types of acquisition methods. In addition, Pulsar is designed to be fast and scalable to large datasets. The false discovery rate (FDR) is controlled at the peptide‑spectrum match (PSM), peptide, and protein‑group level. Furthermore, Pulsar is also able to identify and report co‑fragmented peptides in multiple search rounds by subtracting previously identified fragment ions from the spectra. Pulsar is compatible with data from Thermo Scientific (Orbitrap for both MS1 and MS2) and Sciex, and both centroid and profile modes can be processed.
To generate a library with Pulsar, go to the Prepare Perspective in Spectronaut and click on Generate Library from Pulsar at the bottom left corner (figure 1). A wizard will open to guide you through the process. Briefly, you will be prompted to 1) upload your run files, 2) choose a protein database to perform the search on, 3) the search settings and 4) the library generation settings. To know more details about how to generate a library with Pulsar, refer to the Spectronaut manual here.
Figure 1. Generating a library using on Spectronaut using Pulsar
2. Performing directDIA
To run a directDIA analysis, on the Review Perspective, click on Load directDIA Experiment at the upper left corner (figure 2). Again, a wizard will pop up to guide you setting it up. Briefly, you will be asked to 1) choose your run files, 2) choose your protein database, 3) choose your settings for the search and analysis and 4) provide information about your experimental setup. Optionally, you can also provide gene annotation information to get some biological insight about your experiment.
Figure 2. Setting up a directDIA analysis in Spectronaut. By clicking Next, the wizard will guide you step by step through the process
Created by SEZ. Last update 2018-03-14 by SEZ