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fluorescent-protein-34px-icon.jpg Fluorescent Proteins: Subcellular Localization


Background

The subcellular localization of a protein is often tied to its function, so it's important to determine where your protein of interest resides. There are a variety of methods for determining subcellular localization. One of the most common is to use fluorescence microscopy to compare the localization of your protein of interest with known markers.

Addgene has assembled a collection of fluorescent fusion proteins that are targeted to various organelles or subcellular structures. You should fuse your protein of interest with a fluorescent protein that has a different emission wavelength. For example, if the marker protein is fused to GFP, you may want to fuse your protein to a red fluorescent protein (see this fusion protein guide to find the appropriate backbone). Then, by examining the overlap between your fusion protein and the marker fusion protein using fluorescence microscopy, you can assess whether your protein is targeted to the same structures as these well-characterized proteins.


General Structures

The cell contains a variety of organelles and subcellular structures each with their own unique function. The nucleus, for example, houses the cell’s genetic material. The mitochondria produces the energy cells required for chemical reactions. Labeling these structures can be quite useful for identifying the localization or possible function of a protein of interest. Browse or search the table below to find markers of general cellular organelles and structures.

Plasmid Structure Targeting Gene/Signal Fluorescent Protein PI
pcDNA3.1-kappa-myc-dL5-2xG4S-TMst Cell surface (mammalian) PDGFR-derived dL5 FAP (fluorogen activating protein); mCerulean3 fusion Marcel Bruchez
pcDNA3.1-KozATG-dL5-2XG4S-mCer3 Cytosol, nucleoplasm None dL5 FAP (fluorogen activating protein); mCerulean3 fusion Marcel Bruchez
pcDNA3.1-COXIV-COX8-dL5-2XG4S-mCer3 Mitochondria COX IV-derived (1-22 aa) import sequence and COX VIII signal peptide dL5 FAP (fluorogen activating protein); mCerulean3 fusion Marcel Bruchez
pcDNA3.1-NLS-myc-dL5-2xG4S-mCer3 Nucleus NLS (from Mak16p protein) dL5 FAP (fluorogen activating protein); mCerulean3 fusion Marcel Bruchez
pcDNA3.1-KozATG-myc-dL5-2XG4S-mCer3-SKL Peroxisomes SKL tripeptide, peroxisome transport signal dL5 FAP (fluorogen activating protein); mCerulean3 fusion Marcel Bruchez
pcDNA3.1-kappa-myc-dL5-2XG4S-mCer3-KDEL Endoplasmic reticulum (ER) KDEL dL5 FAP (fluorogen activating protein); mCerulean3 fusion Marcel Bruchez
pHIV-H2BmRFP Chromatin H2B mRFP Bryan Welm, Zena Werb
pEBFP2-Nuc Nucleus NLS EBFP2 Robert Campbell
pmKalama1-Nuc Nucleus NLS mKamala1 Robert Campbell
pCAG-mGFP Membrane palmitoylation sequence from GAP43 EGFP Connie Cepko
pQC NLS mCherry IX Nucleus NLS mCherry Connie Cepko
pQC NLS YFP IX Nucleus NLS YFP Connie Cepko
pQC membrane TdTomato IX Membrane Palmitoylation TdTomato Connie Cepko
FUmGW Membrane Palmitoylation GFP Connie Cepko
pTDpelB-C_sfYFPTwinStrep Periplasmic space PelB signal sequence YFP Thorben Dammeyer
pH2B_mCherry_IRES_puro2 Chromatin H2B mCherry Daniel Gerlich
mCherry-Mito-7 Mitochondria Mitochondrialtargeting sequence(COX8A) mCherry* Michael Davidson
mCherry-LaminB1-10 Nuclear Envelope LaminB1 mCherry* Michael Davidson
mCherry-LaminA-C-18 Nuclear Envelope LaminA1 mCherry* Michael Davidson
mCherry-TOMM20-N-10 Mitochondria-Outer Membrane TOMM20 mCherry* Michael Davidson
mCherry-Peroxisomes-2 Peroxisomes Peroximal Targeting Signal 1 (PTS1) mCherry* Michael Davidson
LV-GFP Chromatin H2B GFP Elaine Fuchs
pCaggs-NLS-PAmCherry1-GSS-EGFP Nucleus NLS mGold Francois St-Pierre
pCMV-mGold-Mito-N-7 Mitochondria Mito mGold Francois St-Pierre
pCMV-mGold-CAF1-C-10 Nucleus CAF-1 mGold Francois St-Pierre
pPalmitoyl-mTurquoise2 Membrane palmitoylation sequence from p63 mTurquoise2 Dorus Gadella
pmTurquoise2-Mito Mitochondria COX8A(1-29) mTurquoise2 Dorus Gadella
pmTurquoise2-H2A Nucleus H2A mTurquoise2 Dorus Gadella
pmTurquoise2-NES Non-nucleus Nuclear Export Sequence mTurquoise2 Dorus Gadella
pmTurquoise2-Peroxi Peroxisomes Peroxisomal Targeting Sequence mTurquoise2 Dorus Gadella
pmScarlet-i_NES_C1 Non-nucleus Nuclear Export Sequence mScarlet-I Dorus Gadella
pmScarlet-H_NES_C1 Non-nucleus Nuclear Export Sequence mScarlet-H Dorus Gadella
pmScarlet_NES_C1 Non-nucleus Nuclear Export Sequence mScarlet Dorus Gadella
pmScarlet-i_H2A_C1 Nucleus H2A mScarlet-I Dorus Gadella
pmScarlet-H_H2A_C1 Nucleus H2A mScarlet-H Dorus Gadella
pmScarlet_H2A_C1 Nucleus H2A mScarlet Dorus Gadella
pmScarlet-I_peroxisome_C1 Peroxisome SRL mScarlet-I Dorus Gadella
pmScarlet-H_peroxisome_C1 Peroxisome SRL mScarlet-H Dorus Gadella
pmScarlet_peroxisome_C1 Peroxisome SRL mScarlet Dorus Gadella
Lck-GFP Membrane palmitoylation sequence from Lck EGFP Steven Green
CAV1-mCherry Caveolae Cav1 mCherry Ari Helenius
pEGFP-C1-Fibrillarin Nucleolus Fibrillarin EGFP Sui Huang
PGK-H2BeGFP Chromatin H2B EGFP Mark Mercola
PGK-H2BmCherry Chromatin H2B mCherry Mark Mercola
pLV-mitoGFP Mitochondria COX8 GFP Pantelis Tsoulfas
mCherry-Parkin Mitochondria Park2 mCherry Richard Youle
H2B-mCherry Chromatin H2B mCherry Robert Benezra
pABCb10-GFP Mitochondria ABCb10 GFP Orian Shirihai
GFP-HP1gamma Chromatin HP1 gamma GFP Tom Misteli
pEGFP-SF2 Nucleus SF2 EGFP Tom Misteli
pLYS1-FLAG-MitoGFP-HA Mitochondria MCU GFP Vamsi Mootha
pH2B-miRFP703 Chromatin H2B miRFP703 Vladislav Verkhusha
pMito-miRFP703 Mitochondria COX8A miRFP703 Vladislav Verkhusha
mito-BFP Mitochondria mitochondrial targeting signal (COX4) TagBFP Gia Voeltz
GFP-Mff Mitochondria-Outer Membrane Mff acGFP1 Gia Voeltz
cfSGFP2-N Extracellular milieu cfSGFP2-N EGFP Ikuo Wada
H2B-GFP Chromatin H2B GFP Geoff Wahl

*Fusions to other FPs can be found by searching the complete Michael Davidson Fluorescent Protein Collection.

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Cytoskeletal Elements

The cytoskeleton is the cell’s internal framework that assists in sustaining structure and function. The cytoskeleton is primarily made up of microtubules, actin filaments, and intermediate filaments. These structures give the cell its shape, organize cellular organelles, and are necessary for movement and cell division. Browse or search the table below to find markers of cytoskeletal element.

Plasmid Structure Targeting Gene/Signal Fluorescent Protein PI
GFP-cortactin Actin Filaments Cortactin EGFP Anna Huttenlocher
pEGFP-N1 alpha-actinin 1 Actin Filaments alpha-actinin 1 EGFP Carol Otey
Cortactin-pmCherryC1 Actin Filaments Cortactin mCherry Christien Merrifield
pLifeAct-mTurquoise2 Actin Filaments LifeAct mTurquoise2 Dorus Gadella
pLifeAct_mScarlet_N1 Actin Filaments LifeAct mScarlet Dorus Gadella
pLifeAct_mScarlet-i_N1 Actin Filaments LifeAct mScarlet-I Dorus Gadella
pLifeAct_mScarlet-H_N1 Actin Filaments LifeAct mScarlet-H Dorus Gadella
pEGFP-C1 F-tractin-EGFP Actin Filaments F-tractin EGFP Dyche Mullins
pCMV-mGold-Actin-C-18 Actin Filaments Actin mGold Francois St-Pierre
mCherry-Lifeact-7 Actin Filaments LifeAct mCherry* Michael Davidson
mEGFP-Lifeact-7 Actin Filaments LifeAct EGFP Michael Davidson
eTC GFP beta-actin full length Actin Filaments beta-actin EGFP Robert Singer
pDEST/LIfeAct-mCherry-N1 Actin Filaments LifeAct mCherry Robin Shaw
pCAG-mGFP-Actin Actin Filaments beta-actin EGFP Ryohei Yasuda
pPAmCherry-b-actin Actin Filaments beta-actin PAmCherry1 Vladislav Verkhusha
pLifeAct-miRFP703 Actin Filaments LifeAct miRFP703 Vladislav Verkhusha
GFP-UtrCH Actin Filaments utrophin (aa# 1-261) GFP William Bement
mCherry-UtrCH Actin Filaments Utrophin mCherry William Bement
GFP-hCCDC11 Centriolar satellites CCDC11 EGFP Ken-Ichi Takemaru
pEGFP Centrin2 Centrioles (dependent on cell cycle) Centrin-2 EGFP Erich Nigg
pEGFP Cep170 C-term Centrioles (dependent on cell cycle) Cep170 EGFP Erich Nigg
pEGFP PICH Centromeres (dependent on cell cycle) PICH EGFP Erich Nigg
mEmerald-PLK1-N-16 Centrosome Plk1 mEmerald* Michael Davidson
pKeratin-miRFP703 Intermediate Filaments (epithelial) Keratin miRFP703 Vladislav Verkhusha
pEGFP-mNFM Intermediate Filaments (neuronal) Nefm EGFP Anthony Brown
pEGFP-rNFM Intermediate Filaments (neuronal) Nefm EGFP Anthony Brown
pVimentin-miRFP703 Intermediate Filaments (vimentin) vimentin miRFP703 Vladislav Verkhusha
pmTurquoise2-Tubulin Microtubules alpha-tubulin mTurquoise2 Dorus Gadella
pmScarlet_alphaTubulin_C1 Microtubules alpha-tubulin mScarlet Dorus Gadella
pmScarlet-i_alphaTubulin_C1 Microtubules alpha-tubulin mScarlet-I Dorus Gadella
pmScarlet-H_alphaTubulin_C1 Microtubules alpha-tubulin mScarlet-H Dorus Gadella
pCMV-mGold-Tubulin-C-18 Microtubules alpha-tubulin mGold Francois St-Pierre
mCh-alpha-tubulin Microtubules alpha-tubulin mCherry Gia Voeltz
pLenti-EB1-EGFP Microtubules EB1 EGFP Ken-Ichi Takemaru
mCherry-Tubulin-C-18 Microtubules alpha-tubulin mCherry* Michael Davidson
pIRESneo-EGFP-alpha Tubulin Microtubules alpha-tubulin EGFP Patricia Wadsworth
pDEST/N1-hEB1-GFP Microtubules EB1 GFP Robin Shaw
pPAmCherry-a-tubulin Microtubules alpha-tubulin PAmCherry Vladislav Verkhusha
pmiRFP703-Tubulin Microtubules alpha-tubulin miRFP703 Vladislav Verkhusha
pEB3-miRFP703 Microtubules EB3 miRFP703 Vladislav Verkhusha
EMTB-3XGFP Microtubules ensconsin GFP William Bement
EMTB-mCherry Microtubules ensconsin mCherry William Bement
GFP-hChibby1 Mother Centrioles / Ciliary Base Chibby1 GFP Ken-Ichi Takemaru
pLenti-EGFP-hChibby1 Mother Centrioles / Ciliary Base Chibby1 EGFP Ken-Ichi Takemaru

*Fusions to other FPs can be found by searching the complete Michael Davidson Fluorescent Protein Collection.

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Cell Junctions

Cell junctions are responsible for forming connections and adhesions between animal cells. They are important for holding cell structures together and reducing cell stress. There are four types of cell junctions: adherens junctions, gap junctions, tight junctions, and desmosomes. Browse or search the table below to find markers for cellular junctions.

Plasmid Structure Targeting Gene/Signal Fluorescent Protein PI
Human Beta-catenin GFP Adherens Junctions Beta-catenin EGFP Alpha Yap
mouse E-cadherin GFP (423) Adherens Junctions E-cadherin EGFP Alpha Yap
Murine E-cadherin mCherry Adherens Junctions E-cadherin mCherry Alpha Yap
mEmerald-Paxillin-22 Focal Adhesions paxillin mEmerald* Michael Davidson
Paxillin-pEGFP Focal Adhesions paxillin GFP Rick Horwitz
RFP-zyxin Focal Adhesions Zyxin RFP Anna_Huttenlocher
pEGFP-N1 alpha-actinin 1 Focal Adhesions alpha-actinin 1 EGFP Carol Otey
pPaxillin-PSmOrange Focal Adhesions paxillin PSmOrange Vladislav Verkhusha
pZyxin-miRFP703 Focal Adhesions Zyxin miRFP703 Vladislav Verkhusha
mCherry-Cx43-7 Gap Junctions Connexin 43 / GJA1-20k mCherry* Michael Davidson
pDEST/hCx43-EGFP-N1 Gap Junctions Connexin 43 EGFP Shaw
pDEST-mCherry-GJA1-20k-N1 Gap Junctions Connexin 43 / GJA1-20k mCherry Shaw
Cx43-msfGFP Gap Junctions Connexin 43 / GJA1 sfGFP David Spray
mApple-VE-Cadherin-N-10 Tight Junctions VE-Cadherin mApple Michael Davidson
mCherry-ZO1-C-14 Tight Junctions Zonula Occludens-1 mCherry Michael Davidson
mCherry-Beta-Catenin-20 Tight Junctions Beta-Catenin mCherry Michael Davidson
pEGFP ZO1 Tight Junctions Zonula Occludens-1 GFP Alan Fanning

*Fusions to other FPs can be found by searching the complete Michael Davidson Fluorescent Protein Collection.

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Secretory Pathway

The secretory pathway is named for its role in secreting proteins from the cell to the extracellular environment. Proteins are first translated at the endoplasmic reticulum, then are transported to the Golgi apparatus in vesicles to be packaged and shipped. Many of the proteins that enter the secretory pathway require chaperones to help with folding or post-translational modification to function properly. Browse or search the table below to find markers of secretory pathway components.

Plasmid Structure Targeting Gene/Signal Fluorescent Protein PI
pEF.myc.ER-E2-Crimson Endoplasmic Reticulum ER retention signal E2-Crimson Glick
pmTurquoise2-ER Endoplasmic Reticulum ER retention signal mTurquoise2 Gadella
mCh-Sec61 beta Endoplasmic Reticulum Sec61 mCherry Voeltz
BFP-Sec61 beta Endoplasmic Reticulum Sec61 BFP Voeltz
BFP-KDEL Endoplasmic Reticulum BiP ER signal sequence BFP Voeltz
Rtn4a-GFP Endoplasmic Reticulum Rtn4a AcGFP Voeltz
ERoxBFP Endoplasmic Reticulum ER retention signal oxBFP Snapp
ERmoxGFP Endoplasmic Reticulum ER retention signal moxGFP Snapp
GolgiCFP Golgi eNOS(1-33) CFP Newton
pmTurquoise2-Golgi Golgi B4GALT1(1-61) mTurquoise2 Gadella
GalToxBFP Golgi GalT signal anchor transmembrane domain oxBFP Snapp
Clathrin-LCa-EYFP Clathrin-coated vesicles Clathrin, light polypeptide EYFP Zhuang
mRFP-Clc Clathrin-coated vesicles Clathrin, light polypeptide mRFP Helenius
mEmerald-Clathrin-15 Clathrin-coated vesicles Clathrin mEmerald* Davidson
DsRed-Rab5 WT Early endosomes RAB5A DsRed2 Pagano
mRFP-Rab5 Early endosomes RAB5A mRFP Helenius
mCh-Rab5 Early endosomes Rab5 mCherry Voeltz
BFP-Rab5 Early endosomes Rab5 BFP Voeltz
GFP-Rab5B Early endosomes Rab5B AcGFP Voeltz
GFP-Rab4B Early endosomes Rab4B AcGFP Voeltz
GFP-rab7 WT Late endosomes RAB7 GFP Pagano
DsRed-rab7 WT Late endosomes RAB7 DsRed Pagano
mCh-Rab7A Late endosomes Rab7a mCherry Voeltz
GFP-Rab7A Late endosomes Rab7a AcGFP Voeltz
GFP-rab11 WT Recycling endosomes RAB11A GFP Pagano
DsRed-rab11 WT Recycling endosomes RAB11A DsRed Pagano
Lamp1-YFP Lysosomes Lamp-1 YFP Mothes
Lamp1-RFP Lysosomes Lamp-1 RFP Mothes
mCherry-Lysosomes-20 Lysosomes LAMP1 mCherry* Davidson
pCMB-AUTr Autophagosome LC3(ATG8) mCherry Harrison
pEGFP-LC3 (human) Autophagosome LC3 EGFP Finkel
pLAMP1-miRFP703 Lysosomes LAMP1 miRFP703 Verkhusha
pmiRFP703-Clathrin Clathrin-coated vesicles Clathrin miRFP703 Verkhusha
pmScarlet-i_Giantin_C1 Golgi apparatus Giantin mScarlet-I Gadella
pmScarlet-H_Giantin_C1 Golgi apparatus Giantin mScarlet-H Gadella
pmScarlet_Giantin_C1 Golgi apparatus Giantin mScarlet Gadella
pCytERM_mScarlet-i_N1 ER CytERM mScarlet-I Gadella
pCytERM_mScarlet-H_N1 ER CytERM mScarlet-H Gadella
pCytERM_mScarlet_N1 ER CytERM mScarlet Gadella
pTag-RFP-C-h-Rab11a-c-Myc Recycling endosomes Rab11a TagRFP Johnson
pTag-BFP-C-h-Rab11a-c-Myc Recycling endosomes Rab11a TagBFP Johnson
pTag-RFP-C-h-Rab7-c-Myc Late endosomes Rab7 TagRFP Johnson
pTag-BFP-C-h-Rab7-c-Myc Late endosomes Rab7 TagBFP Johnson
pTag-RFP-C-h-Rab5a-c-Myc Early endosomes Rab5a TagRFP Johnson
pTag-BFP-C-h-Rab5a-c-Myc Early endosomes Rab5a TagBFP Johnson
pTag-RFP-C-h-Rab4a-c-Myc Recycling endosomes Rab4a TagRFP Johnson
pTag-BFP-C-h-Rab4a-c-Myc Recycling endosomes Rab4a TagBFP Johnson
pcDNA3.1-kappa-myc-dL5-2XG4S-mCer3-KDEL Endoplasmic reticulum (ER) KDEL dL5 FAP (fluorogen activating protein); mCerulean3 fusion Bruchez

*Fusions to other FPs can be found by searching the complete Michael Davidson Fluorescent Protein Collection.

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