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Reference Icon Redbook Plasmids for Optogenetics Research


Background

The field of optogenetics integrates optics and genetic engineering approaches and technology to measure and manipulate cells and their governing biomolecular processes. The tools and technologies developed for optogenetics research utilize light to detect, measure, and control molecular signals, cells, and groups of cells in order to understand their activity and the effects of alterations to this activity.

Optogenetic Schematic

These genetically-encoded tools have come to represent two functions in optogenetics: actuators (genetically-encoded tools for light-activated control of neurons; e.g., microbial opsins) and sensors (genetically-encoded reporters of molecular signals; e.g., calcium indicators). We have organized the optogenetics plasmids available in our repository into actuators and sensors below.

Interested in more news about optogenetics research? Check out the related posts on Addgene's blog.

Plasmids for Optogenetics Actuators

Addgene's repository contains a variety of microbial opsins and additional tools to control neuronal activity. These optogenetics actuators have been organized based on the opsin(s) or tool encoded by the plasmid. Click on the link to jump to plasmids which encode Channelrhodopsins, Halorhodopsins, Archaerhodopsins, Leptosphaeria rhodopsins, or other actuator tools for protein localization, controlled gene expression, and more.

Channelrhodopsins

Variants Description Action
Spectra (nm)
Reporter Plasmid(s)
ChR2 Light-gated cation channel (from Chlamydomonas reinhardtii ) 470 GFP
EYFP
Venus
tdTomato
mCherry
ChR2 mutants ChETA (ChR2 with mutations at position 123) EYFP
tdTomato
ChR2 with various mutations EYFP
mCherry
VChR1 Light-gated cation channel (from Volvox carteri ) 540-570 EYFP
C1C2 ChR1-ChR2 chimera 470 EYFP
ChIEF ChR1-ChR2 chimera with site-directed mutagenesis 470 tdTomato
V1C1 ChR1-VChR1 chimera with E122T and E162T mutations 540 EYFP
mCherry
ChR2-2A-Halo Chr2-GFP and Halo-YFP fused with self-cleaving 2A linker 470/589 GFP/YFP
ChloC SlowChloC: Chloride-conducting cation, ChR2 modification: mutations E90R, D156N, T159C, slow kinetics

ChloC: Chloride-conducting cation, ChR2 modification: mutations E90R, T159C, fast kinetics
465 tdimer2
iChloC Improved ChloC; light-gated chloride channel; AAV backbone; slowChloC mutations plus 2 additional amino acid substitutions (E83Q & E101S) 465 tdimer2
SwiChRca Chloride-conducting cation, iC1C2 modification: mutations C128A 475 (activ.) / 632 (inactiv.) EYFP
Chronos Blue- and green-light drivable (from Stigeoclonium helveticum ) 470, 530 (activ.) GFP
Chrimson Red-light drivable (from Chlamydomonas noctigama ) 590 (activ.) GFP
ChrimsonR K176R mutant of Chrimson 590 (activ.) tdTomato
GFP
GtACR Guillardia theta anion channel rhodopsins
515 EYFP
470

Halorhodopsins

Variants Description Action
Spectra (nm)
Reporter Plasmid(s)
Jaws Red-shifted, light-driven inward chloride pump, used to silence neuronal activity (from Haloarcula (Halobacterium) salinarum (strain Shark); Cruxhalorhodopsin class 632 GFP
Halo (aka NpHR) Light-driven inward chloride pump, used to silence neuronal activity (from Natronomonas pharaonis ) 589 GFP
eNpHR (aka NpHR) Light-driven inward chloride pump, used to silence neuronal activity (from Natronomonas pharaonis ) 589 EYFP
eNpHR 3.0 eNpHR with addition of trafficking signal from Kir2.1 and ER export signal 589 EYFP

Archaerhodopsins

Variants Description Action
Spectra (nm)
Reporter Plasmid(s)
Arch Light-activated outward proton pump (from Halorubrum sodomense ) 566 GFP
CFP
eArch 3.0 Arch with addition of trafficking signal from Kir2.1 566 EYFP
ArchT Light-activated outward proton pump (from Halorubrum genus ) 566 GFP
tdTomato
eArchT 3.0 ArchT with addition of trafficking signal from Kir2.1 566 EYFP

Leptosphaeria rhodopsins

Variants Description Action
Spectra (nm)
Reporter Plasmid(s)
Mac Light-activated outward proton pump (from Leptosphaeria maculans ) 540 GFP
eMac 3.0 Mac with addition of trafficking signal from Kir2.1 540 EYFP

Protein Localization, Controlled Gene Expression, and More

Variants Description Action
Spectra (nm)
Reporter Plasmid(s)
bPAC Light-activated adenylyl cyclase (from Beggiatoa sp.) for light-induced cAMP modulation
BphP1–PpsR2 Near IR induced binding can be used to translocate target proteins to specific cellular compartments or change gene expression. Venus, mCherry
Cryptochrome 2 and CIB1 Blue-light–mediated induction of protein interactions based on Arabidopsis thaliana cryptochrome 2 and CIB1 EGFP, mCherry
Dronpa Light-inducible Dronpa mutant domains that associate and cage a protein in the dark, while dissociate and activate the protein in the light ~400nm (to switch on); ~500nm (to switch off)
iLID Improved light-inducible dimers; Bacterial SsrA peptide is embedded in the LOV2 domain (iLID plasmid), while its natural binding partner SspB is included in a separate plasmid; Blue light activation induces a conformational change in the LOV2 domain which allows SsrA to bind with SspB 450 Venus,
TagRFP-T
Bacterial Mammalian
LAD Light-activated dimerization, using proteins GIGANTEA (GI) and the light oxygen voltage (LOV) domain of FKF1 450
LITEZ Light-inducible transcription using engineered zinc finger proteins, using proteins GIGANTEA (GI) and the light oxygen voltage (LOV) domain of FKF1 450
Lumitoxin Peptide toxin for blocking K + (Kv) channels fused to LOV2 photoswitch mCherry
pDawn Blue-light photoreceptor for light-activated gene expression 470
pDusk Blue-light photoreceptor for light-repressed gene expression 470
Phy–PIF Light-controlled PhyB–PIF6 interaction for reversible translocation of proteins mCherry, mYFP
Rac1-LOV Rac1 fused to photoreactive LOV for localized, reversible Rac activation to control cell motility mCherry, mVenus
TULIPs Tunable light-inducible dimerization tags (TULIPs), plasmids allow cloning of protein coding sequences with GFP-LOVpep, cpPDZ, ePDZb and ePDZb1 as tags 458- or 473-nm GFP, mCherry
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Plasmids for Optogenetics Sensors

Addgene's repository contains a variety of genetically encoded biosensors to monitor and measure fluctuations in molecular signals in cells, including those to monitor calcium, glutamate, pH, and voltage. These optogenetics sensors have been organized below into plasmids containing calcium sensors and other sensors .

Calcium Sensors

Reporter Variant Description Plasmid(s)
GCaMP GCaMP2 Modified calcium indicator derived from a fusion of a cpEGFP with C-terminus CaM and N-terminus M13 (GCaMP)
GCaMP3 Modified GCaMP
GCaMP5G Modified GCaMP
  • GCaMP6s
  • GCaMP6m
  • GCaMP6f
Modified GCaMP
GECO G-GECO1.0 Genetically encoded calcium indicator (GECO)
G-GECO1.1 Modified GECO
G-GECO1.2 Modified GECO
GEM-GECO1 Modified GECO
R-GECO1.0 Modified GECO
GR-GECO1.1 Modified GECO
GR-GECO1.2 Modified GECO
Twitch Twitch-1 High-performing calcium indicator based on C-terminal domain of troponin C
Twitch-2
Twitch-3
Twitch-4
Twitch-5
TN-XXL TN-XXL TN-XXL

Other Sensors

Cellular Signal Reporter Variant Description Plasmid(s)
Chloride & pH ClopHensor Simultaneous, measurement of Cl - and pH with E 2 GFP indicator
ClopHensorN Simultaneous measurement of Cl- and pH optimized for use in the nervous system
Glutamate iGluSnFR Intensity-based glutamate-sensing fluorescent reporter (iGluSnFR)
pH sypHy Fluorescent reporter of pH-sensitive pHluorin fused to synaptophysin
Voltage VSFP VSFP2.1 Voltage-sensitive fluorescent protein (VSFP); Ci-VSP voltage sensor domain fused to a pair of fluorescent proteins (C-terminally truncated CFP and YFP)
VSFP3.1 Modified VSFP with R217Q mutation in the S4 domain of Ci-VSP
VSFP-CR VSFP fused to Clover-mRuby2 FRET pair
ArcLight ArcLight-S249 Modified ArcLight (voltage-sensing domain of Ci-VSP and super ecliptic pHluorin with A227D mutation) with pHluorin at position S249 in Ci-VSP
ArcLight-Q239 Modified ArcLight with pHluorin at position Q239 in Ci-VSP
ArcLight-A242 Modified ArcLight with pHluorin at position A242 in Ci-VSP
ASAP ASAP1 Accelerated Sensor of Action Potentials 1 (ASAP1), a cpGFP is inserted in an extracellular loop of a voltage-sensing domain
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References:

  • Boyden, et al, Nat Neurosci 2005 Sep; 8(9):1263-8.

  • Deisseroth et al., J Neurosci 2006 Oct 11; 26(41):10380-6.

  • Dugué GP et al., Prog Brain Res 2012; 196: 1-28.

  • Alford SC et al., Biol Cell 2013 Jan; 105(1): 14-29.

  • OpenOptogenetics: the optogenetics wiki.

  • Edward Boyden's Lab: Optogenetics Resources

  • Karl Deisseroth's Lab:< a href="http://www.stanford.edu/group/dlab/optogenetics/" target="blank">Optogenetics Resource Center