PFD-rAAV-hSyn-DIO-axon-hChR2(E123T/T159C)-mCherry-WPRE-hGH polyA

rAAV: Recombinant adeno-associated virus. Used as a viral vector, it features high safety (no pathogenicity to humans has been discovered to date), low immunogenicity (rarely eliciting an immune response upon infection), long-term expression, and the ability to infect a wide variety of cells and tissues.

Viral Application: By using the rAAV viral vector to express light-sensitive ion channel proteins (such as ChR2, ChrimsonR, etc.) in neurons, it enables specific excitation of neurons to investigate the effects of optical stimulation on neurons, neural circuits, and animal behavior.

Promoter: hSyn, human synapsin promoter, a mature neuron-specific promoter (448 bp).

Functional Gene: hChR2(E123T/T159C): One of the upgraded versions of ChETA developed in Karl Deisseroth's laboratory. By introducing the T159C point mutation, it significantly improves upon ChETA's drawback of weak inward current; its plateau inward current approaches that of hChR2(H134R) while maintaining ChETA's rapid closing kinetics, endowing the channel with exceptional properties. Building upon the E123T mutation, the T159C mutation increases inward current significantly better than H134R. Excitation light wavelength is 473 nm, suitable for high-frequency optical activation.

Fluorescent Protein: mCherry: Red fluorescent protein, excitation wavelength 587 nm, emission wavelength 610 nm.

Regulatory Element: DIO: Double-floxed Inverse Orientation, an element used in combination with Cre recombinase to control the expression of the target gene. In the absence of Cre recombinase, the target gene is not expressed; normal expression occurs only in the presence of Cre.

Localization Element: axon: Axonal localization.

Notes:

1. Short-term storage at 4°C; long-term storage at -80°C.

2. Can be stored at -80°C for over 6 months. Each freeze-thaw cycle reduces the titer by 10%; repeated freeze-thaw cycles should be avoided. If the virus has been stored for more than 6 months, it is recommended to re-determine the viral titer prior to use.

3. According to experimental requirements, the virus can be appropriately diluted and aliquoted into volumes required for each individual experiment.