AAV with tMCK promoter driven roGFP1

This AAV expresses roGFP1 driven by a muscle tMCK promoter.

tMCK is constructed by ligating a triple tandem copies of mouse MCK enhancer ( about200 bps each) to the ~100bp mouse MCK basal promoter.). This promoter shows extremely sensitive tissue-specificity: (1) In differentiated C2C12 myotubes, the tMCK promoter is 30-50 fold stronger than the Enh358MCK promoter and 10-20 fold stronger than the CMV promoter; (2) In muscle tissue, tMCK promoter is 15-20 fold stronger than the Enh358MCK promoter, and 3-5 fold stronger than the CMV promoter; (3) Strong tissue-specificity: in liver tissue, the expression level from tMCK promoter is only about 0.3-0.5% of that of the CMV promoter.

roGFP1 is a reduction-oxidation sensitive green fluorescent protein used as a biosensor. Two cysteines were introduced into the beta-barrel structure of GFP; the oxidation state, reduced dithiol or the oxidized disulfide, of the engineered thiols determines the fluorescent properties of the sensor. roGFPs (roGFP1 and roGFP2), have two fluorescent excitation maxima at about 400 and 490 nm, respectively and display rapid and reversible ratiometric changes in fluorescence in response to changes in redox potential in vitro and in vivo.