1. Rational design of a monomeric and photostable far-red fluorescent protein for fluorescence imaging in vivo
- Author
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Yuh Nung Jan, Xiaokun Shu, Franck Borel, Dan Yu, Antoine Royant, Matthew P. Klassen, Fanny Marzocca, Rubén Ruiz-González, Su Guo, Zhiqiang Dong, Luca Signor, William A. Weiss, William Clay Gustafson, and Kalpana Makhijani
- Subjects
0301 basic medicine ,Fluorescence-lifetime imaging microscopy ,Biology ,biology.organism_classification ,Biochemistry ,Fluorescence ,Fusion protein ,Photobleaching ,03 medical and health sciences ,030104 developmental biology ,Protein structure ,Centrosome ,Centrin ,Biophysics ,Molecular Biology ,Zebrafish - Abstract
Fluorescent proteins (FPs) are powerful tools for cell and molecular biology. Here based on structural analysis, a blue-shifted mutant of a recently engineered monomeric infrared fluorescent protein (mIFP) has been rationally designed. This variant, named iBlueberry, bears a single mutation that shifts both excitation and emission spectra by approximately 40 nm. Furthermore, iBlueberry is four times more photostable than mIFP, rendering it more advantageous for imaging protein dynamics. By tagging iBlueberry to centrin, it has been demonstrated that the fusion protein labels the centrosome in the developing zebrafish embryo. Together with GFP-labeled nucleus and tdTomato-labeled plasma membrane, time-lapse imaging to visualize the dynamics of centrosomes in radial glia neural progenitors in the intact zebrafish brain has been demonstrated. It is further shown that iBlueberry can be used together with mIFP in two-color protein labeling in living cells and in two-color tumor labeling in mice.
- Published
- 2015