Journal article
A new way to rapidly create functional, fluorescent fusion proteins: random insertion of GFP with an in vitro transposition reaction
BMC neuroscience, Vol.3(1), pp.7-7
06/19/2002
DOI: 10.1186/1471-2202-3-7
PMCID: PMC117241
PMID: 12086589
Abstract
Background: The jellyfish green fluorescent protein (GFP) can be inserted into the middle of another protein to produce a functional, fluorescent fusion protein. Finding permissive sites for insertion, however, can be difficult. Here we describe a transposon-based approach for rapidly creating libraries of GFP fusion proteins.
Results: We tested our approach on the glutamate receptor subunit, GluR1, and the G protein subunit, alpha(s). All of the in-frame GFP insertions produced a fluorescent protein, consistent with the idea that GFP will fold and form a fluorophore when inserted into virtually any domain of another protein. Some of the proteins retained their signaling function, and the random nature of the transposition process revealed permissive sites for insertion that would not have been predicted on the basis of structural or functional models of how that protein works.
Conclusion: This technique should greatly speed the discovery of functional fusion proteins, genetically encodable sensors, and optimized fluorescence resonance energy transfer pairs.
Details
- Title: Subtitle
- A new way to rapidly create functional, fluorescent fusion proteins: random insertion of GFP with an in vitro transposition reaction
- Creators
- Douglas L. Sheridan - Yale UniversityCatherine H. Berlot - Yale UniversityAntoine Robert - Yale UniversityFiona M. Inglis - Yale UniversityKlara B. Jakobsdottir - Yale UniversityJames R. Howe - Yale UniversityThomas E. Hughes - Yale University
- Resource Type
- Journal article
- Publication Details
- BMC neuroscience, Vol.3(1), pp.7-7
- Publisher
- Springer Nature
- DOI
- 10.1186/1471-2202-3-7
- PMID
- 12086589
- PMCID
- PMC117241
- ISSN
- 1471-2202
- eISSN
- 1471-2202
- Number of pages
- 11
- Grant note
- RO1 EY 08362; RO1 GM 50369; RO1 NS 37904 / NIH; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA R01EY008362 / NATIONAL EYE INSTITUTE; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Eye Institute (NEI) R01NS037904 / NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Institute of Neurological Disorders & Stroke (NINDS) R01GM050369 / NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Institute of General Medical Sciences (NIGMS)
- Language
- English
- Date published
- 06/19/2002
- Academic Unit
- Surgery
- Record Identifier
- 9984322767602771
Metrics
23 Record Views