A new reporter gene system suited for cell-free protein synthesis and high-throughput screening in small reaction volumes

The properties of M-hirudin as a new reporter gene system were examined usi ng rabbit reticulocyte lysate for cell-free protein expression. In contrast to the luciferase gene, in vitro translation of M-hirudin is highly robust against changes in concentrations of K+ (and Rb+). In addition, M-hirudin can be detected very sensitively using a reasonably priced fluorimetric thr ombin assay. To show that the new reporter gene system is well suited for ( u)HTS-applications, cell-free synthesis as well as the fluorimetric assay o f M-hirudin were carried out in nanotiter and microtiter plates, respective ly. (C) 2001 Academic Press.