• Rapid establishment of G-protein-coupled receptor-expressing cell lines by site-specific integration.

      Schucht, Roland; Lydford, Simon; Andzinski, Lisa; Zauers, Jeannette; Cooper, James; Hauser, Hansjörg; Wirth, Dagmar; May, Tobias; Department of Gene Regulation and Differentiation, HZI-Helmholtz Centre for Infection Research, Braunschweig, Germany. roland.schucht@inscreenex.com (2011-03)
      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.