Monitoring Protein Splicing Using In-gel Fluorescence Immediately Following SDS-PAGE

Authors

Joel Weinberger II, Murray State University
Christopher W. Lennon, Murray State UniversityFollow

Document Type

Journal Article

Publication Date

8-20-2021

Publication Title

Bio-protocol

Department

Biological Science

College/School

Jesse D. Jones College of Science, Engineering and Technology

Abstract

Inteins garner significant interest from both basic and applied researchers due to their unique catalytic abilities. Herein, we describe a protocol for accurately monitoring protein splicing without purification using in-gel fluorescence immediately following Tris-Glycine SDS-PAGE. Following expression in Escherichia coli, cells are lysed by sonication, cell supernatants are separated using Tris- Glycine SDS-PAGE, and superfolder GFP (sfGFP) fluorescence is directly visualized within gels. This method is rapid, with sfGFP immediately imaged following SDS-PAGE without staining. Further, sfGFP can be specifically detected in complex samples such as E. coli cell supernatants, proteins run at expected masses, and all steps can be performed at ambient temperature. This strategy is broadly applicable beyond the study of protein splicing and can be used for sensitive and specific visualization of superfolder sfGFP-tagged proteins in-gel.

Comments

This is an accepted article published in Bio-protocol, available at https://doi.org/10.21769/BioProtoc.4121

Recommended Citation

Weinberger II, J., & Lennon, C. W. (2021). Monitoring Protein Splicing Using In-gel Fluorescence Immediately Following SDS-PAGE. Bio-protocol, 11(16), e4121-e4121.