Far-red fluorescent tags for protein imaging in living tissues

✍ Scribed by Shcherbo, Dmitry; Murphy, Christopher S.; Ermakova, Galina V.; Solovieva, Elena A.; Chepurnykh, Tatiana V.; Shcheglov, Aleksandr S.; Verkhusha, Vladislav V.; Pletnev, Vladimir Z.; Hazelwood, Kristin L.; Roche, Patrick M.

• Book ID: 111889500
• Publisher: Portland Press
• Year: 2009
• Tongue: English
• Weight: 1013 KB
• Volume: 418
• Category: Article
• ISSN: 0264-6021
• DOI: 10.1042/bj20081949

✦ Synopsis

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.