Stanford University scientists have modified a popular strategy for imaging studies in live animals, making possible a much broader range of biochemical visualization experiments, as described in the April issue of Nature Methods.
The processing of luciferin by the firefly enzyme luciferase generates luminescence, and this luciferase-luciferin system has been used to monitor a wide variety of cellular processes. Helen Blau and her colleagues now introduce a new approach, sequential reporter-enzyme luminescence (SRL), which uses Lugal, a non-toxic and cell-permeable luciferin variant that has been chemically 'caged' so that luciferase cannot process it without the prior action of the enzyme beta-galactosidase (beta-gal). beta-gal is a robust enzyme that can operate in a broad range of conditions, and it rapidly cleaves Lugal to yield a compound that can act as a substrate for luciferase, making luminescence dependent on the expression of both enzymes.
Blau's team show that by injecting Lugal into animals that express luciferase throughout the body but express beta-gal from the promoter of a gene with more tightly regulated expression, they can visually detect changes in that gene's expression over time. With the large number of transgenic mice available expressing beta-gal from different gene promoters, SRL offers a promising tool for the real-time, study of the expression of many genes. The authors also see the possibility that SRL could be adapted for other detection platforms and conclude that "although the system described here uses beta-gal, the concept of SRL should be readily applicable to other commonly used enzymes."
Helen M. Blau (Stanford University School of Medicine, Stanford, CA, USA)
Abstract available online.
(C)Nature Methods press release.
Message posted by: Trevor M. D'Souza
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