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. 2021 Mar;1(3):e69.
doi: 10.1002/cpz1.69.

PIONEER: Pipeline for Generating High-Quality Spectral Libraries for DIA-MS Data

Srikanth S Manda  1 , Zainab Noor  1 , Peter G Hains  1 , Qing Zhong  1
Affiliations

PIONEER: Pipeline for Generating High-Quality Spectral Libraries for DIA-MS Data

Srikanth S Manda et al. Curr Protoc. 2021 Mar.

Erratum in

Abstract

Data-independent-acquisition mass spectrometry (DIA-MS) is a state-of-the-art proteomic technique for high-throughput identification and quantification of peptides and proteins. Interpretation of DIA-MS data relies on the use of a spectral library, which is optimally created from data acquired from the same samples in data-dependent acquisition (DDA) mode. As DIA-MS quantification relies on the spectral libraries, having a high-quality, non-redundant, and comprehensive spectral library is essential. This article describes the major steps for creating a high-quality spectral library using a combination of multiple complementary search engines. We discuss appropriate strategies to control the false discovery rate for the final spectral library as a result of merging multiple searches. © 2021 The Authors Current Protocols © 2021 Wiley Periodicals LLC. Basic Protocol 1: Searching DDA-MS files with multiple search engines Basic Protocol 2: Merging results from multiple search engines Basic Protocol 3: Creating spectral libraries from merged results Alternate Protocol: Using CLI for automating tasks Support Protocol: Creating concatenated FASTA files.

Keywords: DIA; SWATH; mass spectrometry; proteomics; spectral library.

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References

Literature Cited

References
    1. Barkovits, K., Pacharra, S., Pfeiffer, K., Steinbach, S., Eisenacher, M., Marcus, K., & Uszkoreit, J. (2020). Reproducibility, specificity and accuracy of relative quantification using spectral library-based data-independent acquisition. Molecular and Cellular Proteomics, 19(1), 181-197. doi: 10.1074/mcp.RA119.001714
    1. Barsnes, H., & Vaudel, M. (2018). SearchGUI: A highly adaptable common interface for proteomics search and de novo engines. Journal of Proteome Research, 17(7), 2552-2555. doi: 10.1021/acs.jproteome.8b00175
    1. Bruderer, R., Bernhardt, O. M., Gandhi, T., Miladinovic, S. M., Cheng, L. Y., Messner, S., … Reiter, L. (2015). Extending the limits of quantitative proteome profiling with data-independent acquisition and application to acetaminophen-treated three-dimensional liver microtissues. Molecular and Cellular Proteomics, 14(5), 1400-1410. doi: 10.1074/mcp.M114.044305
    1. Chambers, M. C., Maclean, B., Burke, R., Amodei, D., Ruderman, D. L., Neumann, S., … Mallick, P. (2012). A cross-platform toolkit for mass spectrometry and proteomics. Nature Biotechnology, 30(10), 918-920. doi: 10.1038/nbt.2377
    1. Cho, J. Y., Lee, H. J., Jeong, S. K., Kim, K. Y., Kwon, K. H., Yoo, J. S., … Paik, Y. K. (2015). Combination of multiple spectral libraries improves the current search methods used to identify missing proteins in the chromosome-centric human proteome project. Journal of Proteome Research, 14(12), 4959-4966. doi: 10.1021/acs.jproteome.5b00578

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