A vast colour palette of monomeric fluorescent proteins has been developed to investigate protein localization, motility and interactions. However, low brightness has remained a problem in far-red variants, which hampers multicolour labelling and whole-body imaging techniques. In the present paper, we report mKate2, a monomeric far-red fluorescent protein that is almost 3-fold brighter than the previously reported mKate and is 10-fold brighter than mPlum. The high-brightness, far-red emission spectrum, excellent pH resistance and photostability, coupled with low toxicity demonstrated in transgenic Xenopus laevis embryos, make mKate2 a superior fluorescent tag for imaging in living tissues. We also report tdKatushka2, a tandem far-red tag that performs well in fusions, provides 4-fold brighter near-IR fluorescence compared with mRaspberry or mCherry, and is 20-fold brighter than mPlum. Together, monomeric mKate2 and pseudo-monomeric tdKatushka2 represent the next generation of extra-bright far-red fluorescent probes offering novel possibilities for fluorescent imaging of proteins in living cells and animals.
Far-red fluorescent tags for protein imaging in living tissues
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Dmitry Shcherbo, Christopher S. Murphy, Galina V. Ermakova, Elena A. Solovieva, Tatiana V. Chepurnykh, Aleksandr S. Shcheglov, Vladislav V. Verkhusha, Vladimir Z. Pletnev, Kristin L. Hazelwood, Patrick M. Roche, Sergey Lukyanov, Andrey G. Zaraisky, Michael W. Davidson, Dmitriy M. Chudakov; Far-red fluorescent tags for protein imaging in living tissues. Biochem J 15 March 2009; 418 (3): 567–574. doi: https://doi.org/10.1042/BJ20081949
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