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Red-shifted Renilla reniformis luciferase variants for imaging in living subjects

Abstract

The use of R. reniformis luciferase (RLuc) as a reporter gene in small-animal imaging has been hampered by its 481 nm peaked emission spectrum, as blue wavelengths are strongly attenuated in biological tissues. To overcome this, we generated variants of RLuc with bathochromic (red) shifts of up to 66 nm (547 nm peak) that also had greater stability and higher light emission than native RLuc.

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Figure 1: Active site residues and the effects of mutations on the emission spectrum of RLuc.
Figure 2: Testing RLuc8.6-535 in cell culture and in mice.

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Acknowledgements

This work was supported in part by a Stanford Bio-X Graduate Fellowship (A.M.L.), US National Cancer Institute (NCI) CA114747 ICMIC P50 (S.S.G.), NCI CA119367 CCNE U54 (SSG), NCI R01 CA082214 (SSG), and the NCI Small Animal Imaging Resource Program. We thank B. Rice and T. Troy of Caliper Life Sciences for help with the spectra measurements.

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Correspondence to Sanjiv Sam Gambhir.

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Stanford University has filed a patent application that includes portions of the research described in this manuscript.

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Supplementary Figures 1–4, Supplementary Tables 1–3, Supplementary Methods. (PDF 933 kb)

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Loening, A., Wu, A. & Gambhir, S. Red-shifted Renilla reniformis luciferase variants for imaging in living subjects. Nat Methods 4, 641–643 (2007). https://doi.org/10.1038/nmeth1070

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