A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
Summary: Some protein-protein interactions are induced by different kinds of stimulation and are dependent on specific genes. To identify these interaction partners, we present a protocol which utilizes affinity purification of Flag-tagged protein complexes followed by mass-spectrometry-based proteo...
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| Main Authors: | , |
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| Format: | article |
| Language: | EN |
| Published: |
Elsevier
2021
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| Subjects: | |
| Online Access: | https://doaj.org/article/5b067d16d3214e3b9d2f7d55f45809db |
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| Summary: | Summary: Some protein-protein interactions are induced by different kinds of stimulation and are dependent on specific genes. To identify these interaction partners, we present a protocol which utilizes affinity purification of Flag-tagged protein complexes followed by mass-spectrometry-based proteomics to compare stimulation-induced interactomes between wild-type and CRISPR-Cas9-mediated knockout cells. The candidates of interest are identified using bioinformatic analyses and verified by biochemical approaches. This protocol is highly versatile and applies to a variety of cells and different types of stimulation.For complete details on the use and execution of this protocol, please refer to (Zhu et al., 2021). |
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