Optogenetics
rAAV-CaMKIIa-DIO-OChIEF-P2A-dTomato-WPRE-bGH polyA
The excitatory neuron-specific promoter CaMKIIa regulates the expression of Cre-dependent light-activated elements and dTomato. OChIEF-dTomato is expressed in the presence of Cre, but OChIEF-dTomato is not expressed in the absence of Cre. OChIEF and dTomato are connected with 2A and expressed separately.
AAV1, AAV2, AAV5, AAV6, AAV8, AAV9, rh10, DJ, B10, PHP.eB, PHP.S, MaCPNS1/2, retro, anc80l65, Please consult for more serotypes
OChIEF(E163A/T199C)
CaMKIIa
Store at -80°C
Delivery Information
Usually Ships in 7 Business Days；
Product Details
Promoter Details
CaMKIIa, pyramidal neuron/excitatory neuron promoter, 1293bp.
Functional Gene Details
OChIEF: ChIEF is a fusion of two proteins, ChR1 and ChR2, with Ile mutated to Val at position 170. The characteristic of this protein is that under long-term illumination, the ratio of plateau current to peak current is large, which means that the channel is less inactivated under illumination and is suitable for long-term stimulation. The mutation of Ile accelerates the closing speed of the channel, making ChIEF suitable for high-frequency stimulation. Karl's laboratory has compared the high-frequency stimulation of ChIEF and hChR2(E123A) in pv (small serum protein) neurons, and the effects are equivalent. OChIEF is a codon-optimized version. The excitation light wavelength is 473nm, suitable for high-frequency light activation.
Fluorescent Tag
dTomato
Fluorescent Protein Details
dTomato: Orange fluorescent protein, excitation wavelength 554nm, emission wavelength 581nm.
Virus Type
adeno-associated virus
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.
Regulatory Element Details
DIO: Double-floxed inverse Orientation, is an element that cooperates with Cre recombinase to control the expression of the target gene. If there is no Cre recombinase, the target gene will not be expressed, but in the presence of Cre, the target gene can be expressed normally. 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.
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.
