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. 2010 Jan;7(1):5-6.
doi: 10.1038/nmeth0110-05.

A red-shifted Renilla luciferase for transient reporter-gene expression

Andreas Markus Loening, Anca Dragulescu-Andrasi, Sanjiv Sam Gambhir

A red-shifted Renilla luciferase for transient reporter-gene expression

Andreas Markus Loening et al. Nat Methods. 2010 Jan.
No abstract available

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Conflict of interest statement

Competing Financial Interests

The authors declare competing financial interests.

Figures

Figure 1
Figure 1
Validation of RLuc7-521 (a) Normalized bioluminescence emission spectra for RLuc and RLuc7-521 in the presence of coelenterazine (c) or coelenterazine-v (v). The peaks for RLuc were 481 nm (c) and 516 nm (v), with mean emission wavelengths of 500 nm (c) and 535 nm (v). The peaks for RLuc7-521 were 521 nm (c) and 556 nm (v), with mean emission wavelengths of 534 nm (c) and 569 nm (v). Normalization in the figure is to the total area under the curve, not the emission peak. (b) Mammalian cell expression of RLuc and RLuc7-521 following transient transfection into 293T cells. The luciferases were in pcDNA 3.1 plasmids under the control of the constitutive promoter from cytomegalovirus (CMV). Cells were exposed to 100 μg/ml of cycloheximide to inhibit new protein synthesis 24 hours following transfection. Cells were assayed for light out-put per total cellular protein. Data was fit to a mono-exponential degradation model and estimated intracellular activity half-lives are given in the figure key. Values are reported as relative to the RLuc condition immediately prior to cycloheximide application (T=0). The two conditions were significantly different (p <0.05, two-tailed Student t-test) at all time points except the last. Samples were in quadruplicate, error bars represent standard error of the mean. (c) Bioluminescence from 293T cells containing either RLuc or RLuc7-521 within the lungs of nude mice. Cells were transiently transfected with pcDNA 3.1 plasmids containing either RLuc or RLuc7-521, 24 h later these cells were injected via the tail-vein with resultant accumulation in the lungs. Mice were injected with coelenterazine 2 h later and imaged. Representative mice from a substrate control group (n=5), the RLuc condition (n=7), and the RLuc7-521 condition (n=7) are shown. Total flux from the thorax was recorded, with substrate-only injected mice used for background subtraction. As shown in the bar graph, the two conditions were significantly different (p <0.001, two-tailed Student t-test).
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References

    1. Zhao H, et al. J Biomed Opt. 2005;10:41210. - PubMed
    1. Loening AM, Wu AM, Gambhir SS. Nat Methods. 2007;4:641–643. - PubMed
    1. Inouye S, Shimomura O. Biochem Biophys Res Commun. 1997;233:349–353. - PubMed
    1. Loening AM, Fenn TD, Wu AM, Gambhir SS. Protein Eng Des Sel. 2006;19:391–400. - PubMed
    1. Loening AM, Fenn TD, Gambhir SS. J Mol Biol. 2007;374:1017–1028. - PMC - PubMed

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