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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.
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Figure. Arabidopsis seedlings expressing MatryoshCaMP6s report root cytosolic calcium elevation in response to salt shock.

  

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*2018/10/08 10:30 AM
Dr. Bernhard Grimm (Dean of the Life Sciences Faculty, Professor (C4) and Head of Department of Plant Physiology at the Institute of Biology, Humboldt-Universität Berlin, Germany)
Green or red? Synthesis of chlorophyll and heme - Multiple posttranslational control of glutamyl-tRNA reductase, the rate limiting enzyme of tetrapyrrole biosynthesis
Auditorium A134, Agricultural Technology Building, Agricultural Biotechnology Research Center

*2018/10/29 10:30 AM
Dr. Ling Li (Assistant Professor, Department of Biological Sciences, Mississippi State University, USA)
A molecular tool to increase protein content and broad disease resistance in crops
Auditorium A134, Agricultural Technology Building, Agricultural Biotechnology Research Center

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