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

J Biomol Screen. 2011 Mar;16(3):323-31. doi: 10.1177/1087057110396371. Epub 2011 Feb 18.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CHO Cells
  • Cell Line
  • Cricetinae
  • Cricetulus
  • Gene Expression Regulation
  • Gene Order
  • Gene Targeting*
  • Genetic Vectors
  • High-Throughput Screening Assays
  • Receptors, G-Protein-Coupled / genetics*
  • Receptors, G-Protein-Coupled / metabolism*
  • Recombinases / metabolism

Substances

  • Receptors, G-Protein-Coupled
  • Recombinases