A bright monomeric green fluorescent protein derived from Branchiostoma lanceolatum

Abstract

We report a monomeric yellow-green fluorescent protein, mNeonGreen, derived from a tetrameric fluorescent protein from the cephalochordate Branchiostoma lanceolatum. mNeonGreen is the brightest monomeric green or yellow fluorescent protein yet described to our knowledge, performs exceptionally well as a fusion tag for traditional imaging as well as stochastic single-molecule superresolution imaging and is an excellent fluorescence resonance energy transfer (FRET) acceptor for the newest cyan fluorescent proteins.

Figure 1

Fluorescence imaging of mNeonGreen (mNG) fusion vectors. C-terminal mNG fusion constructs (with respect to the fluorescent protein); for each fusion, the linker amino acid length is indicated after the name of the targeted organelle or fusion partner: (a) mNG-Annexin (A4)-C-12 (human; plasma membrane); (b) mNG-β-actin-C-18 (human; actin cytoskeleton); (c) mNG-β-Catenin-C-20 (mouse; tight junctions); (d) mNG-CAAX-C-5 (20-amino acid farnesylation signal from c-Ha-Ras; plasma membrane); (e) mNG-CAF1-C-10 (mouse chromatin assembly factor); (f) mNG-Caveolin1-C-10 (human); (g) mNG-Endosomes-C-14 (human RhoB GTPase); (h) mNG-Fascin-C-10 (human; actin bundling); (i) mNG-Fibrillarin-C-7 (human; nucleoli); (j) mNG-FilaminA-C-14 (human; cytoskeleton); (k) mNG-LAMP1-C-20 (rat; lysosomal membrane glycoprotein 1; lysosomes); (l) mNG-Clathrin-C-15 (human, light chain B); (m) mNG-Myotilin-C-14 (human; actin filaments); (n) mNG-PCNA-C-19 (human; replication foci); (o) mNG-Plastin-C-10 (human; actin binding); (p) mNG-Rab4a-C-7 (human; endosomes); (q) mNG-LC3B-C-7 (rat light chain; autophagosoms; (r) mNG-Talin-C-18 (mouse; focal adhesions); (s) mNG-Tubulin-C-35 (human; microtubules); (t) mNG-ZO1-C-14 (human; tight junctions). The cell line used for expression of C-terminal mNG constructs was Madin-Darby canine kidney (MDCK; ATCC, CCL-34) cells in panels (c) and (t). HeLa CCL2 (ATCC) cells were used in the remaining panels. Scale bars represent 10 μm.