Transient expression of DNA inDrosophilavia electroporation

The capability of introducing DNA into Drosophila embryos using an electric field pulse is easier and more efficient than current procedures involving microinjection (1). With short electric impulses above a certain field strength, biomembranes are made transiently more permeable which leads to an exchange of matter across the perturbed membrane (2). Drosophila embryos, which are incubated in a solution of DNA and exposed to an electric field pulse can take up and transiently express this DNA. To assess the transient expression of DNA, we measured aldehyde oxidase (AO) activity, which is normally expressed in a tissue specific manner (Figure 1A) (3). AO activity depends upon the sulfurated form of a molybdenum cofactor. Sulfuration of the cofactor requires the maroon-like (ma-l) gene; ma-l mutants show complete absence of AO activity (Figure IB) (4). Under optimized conditions (960 /xF capacitor at 250 V/cm, 10 /tg/ml ma-l phage DNA), we ascertained that with non-dechorionated embryos 75% survived to 3rd instar larvae, and 95% of these had taken up functional ma-l DNA. Expression levels were equivalent to that attainable with microinjection (5), and exhibit the typical mosaic staining pattern shown in Figure 1C. To achieve nearly wild type expression levels, embryos were dechorionated prior to electroporation. Optimized conditions (960 pF capacitor at 70 V/cm, 10 /tg/ml ma-l phage DNA) resulted in 10% of embryos surviving to 3rd instar larvae and of those surviving, 100% had taken up functional ma-l DNA (Figure ID). The extraordinary ease with which hundreds of embryos may be electroporated makes this the method of choice for transient expression. Since transient expression is controlled by adjacent promoter/enhancer elements, the nearly wild type expression level in most surviving larvae suggests that electroporation can be a powerful means for studying such regulatory regions without the necessity for germ-line transformation. Moreover, high levels of transient expression suggest that the rescue of developmental lethality may be possible in some cases. Since electroporation of dechorionated embryos also results in expression of AO activity in posterior structures (data not shown), experiments attempting germ-line transformation of Drosophila by electroporation are underway.