AAV with hRPE(0.8) promoter driven FLPo

This AAV expresses FLPo driven by a retina hRPE(0.8) promoter.

The RPE65 gene encodes a protein of the retinal pigment epithelium (RPE) with a key role in the cycling of retinoids. It was shown that a RPE promoter of about 0.8 Kb was sufficient to direct high RPE-specific expression in transgenic mice. The 1.5Kb version of the promoter hRPE(1.5), also gives RPE-specific expression, but the expression level is not as high as the 0.8Kb promoter. This is probably due to a negative regulatory element(s) present in the 1.5Kb promoter.

FLPo is a codon-optimized version of FLPe that greatly increases the protein expression and the FRT recombination efficiency in mouse cells. FLP is a site-specific recombinase (SSR) from the _ integrase family which recognizes distinct 34 bp FRT sites. Like Cre/LoxP system, the FLP/FRT system has been widely used for gene expression and generating conditional knockout mice, mediated by the FLP/FRT system. Initial use of FLP in mammalian cells revealed inefficient recombinase activity due to thermal instability of the FLP protein. Subsequent screening for thermostable mutants resulted in the identification of FLPe which has a 4-fold higher recombination efficiency at 37oC than original FLP. However, the recombination efficiency of FLPe in cells remains very low relative to other recombinases because of its non-mammalian origin. FLPo is engineered by codon optimization of FLPe, and gives much higher expression and activity than FLPe.