Using Inteins to Label Proteins with GFP
Author Information
Author(s): Richard Ramsden, Luther Arms, Trisha N. Davis, Eric G. D. Muller
Primary Institution: University of Washington
Hypothesis
Can inteins be modified to incorporate selectable genetic markers while maintaining splicing efficiency?
Conclusion
The Pch PRP8 intein can incorporate various genetic markers and still exhibit high splicing activity, allowing for the insertion of GFP into target proteins in yeast.
Supporting Evidence
- The modified inteins spliced at high efficiency in both E. coli and yeast.
- Splicing efficiency was greater than 96% when expressed from a plasmid in S. cerevisiae.
- The His5-marked intein yielded a fully functional calmodulin tagged with GFP.
Takeaway
Inteins are special proteins that can cut themselves out of larger proteins. This study shows how we can use them to add a green glow to other proteins easily.
Methodology
The study involved modifying the Pch PRP8 intein to include selectable genetic markers and testing splicing efficiency in both E. coli and yeast.
Limitations
The G418R and HygR inteins showed impaired growth and inefficient splicing when integrated into the CMD1 locus.
Digital Object Identifier (DOI)
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