Abstract
The establishment of mammalian cell lines reliably expressing G-protein-coupled receptors (GPCRs) can be a tedious and often time-consuming process. A strategy has been developed to allow the rapid production of such cell lines. The first step of this approach was the generation of a specialized master cell line, characterized by optimized stable expression of a membrane-bound reporter protein. In the second step, this reporter gene was exchanged for that of the GPCR of interest by a DNA recombinase "cut-and-paste" engineering step. It has been demonstrated that the resulting GPCR cell lines inherit the advantages of the master cell line, expressing the GPCR in a homogeneous and stable manner. The case studies presented demonstrate the functionality of the established GPCR cell lines, and most important, because of the highly efficient integration event, these recombinant GPCR-expressing cell lines were generated within a timeframe of 2 to 4 weeks. The advantages of this cut-and-paste approach versus other strategies such as Flp-In or Jump-In are compared. (Journal of Biomolecular Screening 2011;16:323-331)
| Original language | English |
|---|---|
| Pages (from-to) | 323-331 |
| Number of pages | 9 |
| Journal | Journal of Biomolecular Screening |
| Volume | 16 |
| Issue number | 3 |
| DOIs | |
| Publication status | Published - Mar 2011 |
| Externally published | Yes |
Bibliographical note
Funding Information:This work was supported by grants from the Helmholtz Society (Helmholtz Enterprise Fund), by the Niedersächsisches Ministerium für Wissenschaft und Kultur (80029155), and by grants of the German Ministry for Research and Education (FKZ 0313940 and PTJ-Bio/0313735/Nn07-06) and the Deutsche Forschungsgemeinschaft (Excellence Cluster REBIRTH).
Keywords
- Flp-In
- G-protein-coupled receptors
- Jump-In
- cell-based assays
- gene expression
- recombinase mediated cassette exchange