PFD-rAAV-hSyn-hChR2(H134R)-P2A-eNpHR3.0-EGFP-WPREs

Promoter Details

hSyn, human synapsin promoter, mature neuron-specific promoter, 448bp.

Functional Gene Details

hChR2 (H134R): It is the most common form of ChR2. h refers to the coding sequence of the gene that has been optimized for the human codon table. H134R refers to the mutation of histidine at position 134 to arginine. This mutation inhibits the binding of H+ by ChR2 and increases the passage ability of Na+, thereby increasing the overall conductivity of the molecule and thereby increasing the inward current excited by light. Generally, blue light around 470nm is used for excitation. eNpHR3.0: NpHR is a light-driven chloride ion pump belonging to the Halorhodopsin family. The common version is NpHR3.0, which adds an endoplasmic reticulum anterograde transport signal and a Golgi transport signal to the cell membrane based on NpHR, improving its expression on the cell membrane. It is the most commonly used photoinhibition tool. NpHR is usually driven by yellow light around 570nm and can be used simultaneously with hChR2 (H134R).

eNpHR3.0: NpHR is a light-driven chloride ion pump belonging to the Halorhodopsin family. The common version is NpHR3.0, which adds an endoplasmic reticulum anterograde transport signal and a Golgi transport signal to the cell membrane based on NpHR, improving its expression on the cell membrane. It is the most commonly used photoinhibition tool. NpHR is usually driven by yellow light around 570nm and can be used simultaneously with hChR2 (H134R). hChR2 (H134R): It is the most common form of ChR2. h refers to the coding sequence of the gene that has been optimized for the human codon table. H134R refers to the mutation of histidine at position 134 to arginine. This mutation inhibits the binding of H+ by ChR2 and increases the passage ability of Na+, thereby increasing the overall conductivity of the molecule and thereby increasing the inward current excited by light. Generally, blue light around 470nm is used for excitation.

hChR2 (H134R): It is the most common form of ChR2. h refers to the coding sequence of the gene that has been optimized for the human codon table. H134R refers to the mutation of histidine at position 134 to arginine. This mutation inhibits the binding of H+ by ChR2 and increases the passage ability of Na+, thereby increasing the overall conductivity of the molecule and thereby increasing the inward current excited by light. Generally, blue light around 470nm is used for excitation.

eNpHR3.0: NpHR is a light-driven chloride ion pump belonging to the Halorhodopsin family. The common version is NpHR3.0, which adds an endoplasmic reticulum anterograde transport signal and a Golgi transport signal to the cell membrane based on NpHR, improving its expression on the cell membrane. It is the most commonly used photoinhibition tool. NpHR is usually driven by yellow light around 570nm and can be used simultaneously with hChR2 (H134R).

Fluorescent Tag

EGFP

Fluorescent Protein Details

EGFP: Green fluorescent protein, excitation wavelength 488nm, emission wavelength 507nm. 2A: The 2A sequence is a short peptide sequence. When the ribosome translates it, it will automatically jump away and continue to translate the downstream reading frame, thereby translating the same reading frame into two independent proteins. Used in non-fusion expressions. Commonly used 2A sequences include T2A and P2A.

EGFP: Green fluorescent protein, excitation wavelength 488nm, emission wavelength 507nm. 2A: The 2A sequence is a short peptide sequence. When the ribosome translates it, it will automatically jump away and continue to translate the downstream reading frame, thereby translating the same reading frame into two independent proteins. Commonly used 2A sequences include T2A and P2A. Used in non-fusion expressions.

EGFP: Green fluorescent protein, excitation wavelength 488nm, emission wavelength 507nm.

Application

Expressing light-sensitive ion channel proteins, such as ChR2, ChrimsonR, etc., in neurons through rAAV viral vectors can specifically excite neurons and explore the effects of light stimulation on neurons, neural circuits and animal behavior.

Expressing light-sensitive ion channel proteins, such as eNpHR3.0, Arch, etc., in neurons through rAAV viral vectors can specifically inhibit neurons and explore the effects of light stimulation on neurons, neural circuits and animal behavior.

Notes

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

2. It can be stored at -80°C for more than 6 months. Each time it is frozen and thawed, the titer will be reduced by 10%, and repeated freezing and thawing should be avoided; if the virus is stored for more than 6 months, it is recommended to re-measure the virus titer before use

3. According to the experimental needs, the virus can be appropriately diluted and divided into the volume required for each experiment.