The gypsy insulator of Drosophila melanogaster, together with its binding protein suppressor of Hairy-wing, facilitate high and precise expression of transgenes in Arabidopsis thaliana.

Abstract

The variation of expression pattern exhibited by a transgene as a result of random integration, known as position effect, is, among other mechanisms, a particular challenge to reverse genetics. We present a strategy to counteract position effect in Arabidopsis thaliana by flanking the transgenes with the gypsy insulator from Drosophila melanogaster. In addition, Suppressor of Hairy-wing [Su(Hw)], the binding protein of the gypsy insulator, was coexpressed. Results indicated that the gypsy insulators could efficiently improve the expression levels of reporter genes driven by various kinds of promoters by 8- to 13-fold. Coexpression of the Su(Hw) protein led to a more uniform expression level of transgenes, as the coefficient of variation of expression levels was reduced further. The gypsy-Su(Hw) system enhanced expression levels, but did not alter the specificity of promoter activities, as experimentally evidenced by the promoters of the PIN and the AFB gene families. Interestingly, the gypsy insulator was also able to improve the expression of a selectable marker gene outside the insulated region, which facilitated the screen of transformants. Our system will likely decrease the number of lines that experimenters need to create and examine for a given transgene by contributing to relatively high and precise expression of transgenes in plants. Certain features of the gypsy insulator in Arabidopsis also provide new perspectives on the insulator field.