Cindy Ast, Jessica Foret, Luke M. Oltrogge, Roberto De Michele, Thomas J. Kleist, Cheng-Hsun Ho & Wolf B. Frommer (2017) Ratiometric Matryoshka biosensors from a nested cassette of green- and orange-emitting fluorescent proteins.Nature Communications(2017), doi:10.1038/s41467-017-00400-2
Sensitivity, dynamic and detection range as well as exclusion of expression and instrumental artifacts are critical for the quantitation of data obtained with fluorescent protein (FP)-based biosensors in vivo. Current biosensors designs are, in general, unable to simultaneously meet all these criteria. Here, we describe a generalizable platform to create dual-FP biosensors with large dynamic ranges by employing a single FP-cassette, named GO-(Green-Orange) Matryoshka. The cassette nests a stable reference FP (large Stokes shift LSSmOrange) within a reporter FP (circularly permuted green FP). GO- Matryoshka yields green and orange fluorescence upon blue excitation. As proof of concept, we converted existing, single-emission biosensors into a series of ratiometric calcium sensors (MatryoshCaMP6s) and ammonium transport activity sensors (AmTryoshka1;3). We additionally identified the internal acid-base equilibrium as a key determinant of the GCaMP dynamic range. Matryoshka technology promises flexibility in the design of a wide spectrum of ratiometric biosensors and expanded in vivo applications.
Figure. Arabidopsis seedlings expressing MatryoshCaMP6s report root cytosolic calcium elevation in response to salt shock.
2018/03/19 10:00 AM
Dr. Masashi Yamada (Postdoctoral Fellow, Duke University, U.S.A.)
Function of alternatively spliced isoforms and a small peptide in root development
Auditorium A134, Agricultural Technology Building, Agricultural Biotechnology Research Center 2018/04/09 10:30 AM
Dr. Suh-Chin Wu (Professor, Institute of Biotechnology, National Tsing Hua University, Taiwan)
Rational design of site-specific glycan-masking and glycan-unmasking HA antigens as new-generation influenza vaccines
Auditorium A134, Agricultural Technology Building, Agricultural Biotechnology Research Center