AAV with CMV7 promoter driven Cre Inducible hChR2(C128T)-mCherry

This AAV expresses DIO-hChR2(C128T)-mCherry driven by an ubiquitous CMV7 promoter.

The CMV7 promoter is about 0.35 kb in length and is a shortened version of the commonly used CMV promoter. CMV7 has a truncated version of the CMV enhancer. When tested in several cell lines, this promoter provides an expression level that is about 3-5 fold lower than that of the original 0.6Kb CMV promoter.

hChR2 is a humanized version of ChR2 for mammalian expression. Wild-type ChR2, as well as a few of its mutations, provides the fastest excitation of the channelrhodopsins offered, and is widely used in in optogenetics techniques in neuroscience. Mutations at C128 (C128A, C128S, C128T) can extend depolarization up to seconds or even minutes; they are sometimes referred to as step-function opsins or SFOs. hChR2(C128T) mutant provides for a 2-second open-state lifetime. It has showed strong expression in the hippocampal neurons and Pichia pastoris. It becomes inactivated at strong light due to a reaction into a deprotonated species.

In the DIO scenario, the transgene of interest is inserted in reverse orientation relative to the 5' promoter and is flanked by oppositely oriented loxP and lox2272 sites. In the absence of Cre expression, the transgene will not be produced. In the presence of Cre expression, the transgene will be "FLip-EXchanged" or FLEXed, leading to expression of the transgene. This is due to a permanent Cre-mediated recombination/inversion of the flanked transgene. This arrangement is called DIO (double-floxed inverse ORF), Cre-ON, Flex-rev (reverse), Flex-ON/FlexON, or DIO-AAV/AAV-DIO (double-floxed inverse ORF in AAV).