关键词: 黑腹果蝇, Gal4/UAS系统,  Gal80^ts, UAS转基因, 基因表达水平, 荧光强度

Abstract: 【Aim】 The combination of Gal80^ts and Gal4 is commonly used to regulate the UAS transgene overexpression in Drosophila, and flexibly turns on or off the transgene expression in a temperature-dependent manner. Gal80^ts, a temperature-sensitive protein, binds Gal4 and inhibits its transcription activity at low temperature (18℃), but relieves the inhibition of Gal4 protein and allows its binding to the UAS sites to drive the expression of UAS transgenes at high temperature (29℃). However, the switch from 18 to 29℃ could only achieve intensive overexpression of UAS transgenes, but not flexible manipulation. Thus in this study we systematically studied the flexible control of UAS transgene expression levels by this system at a series of temperatures. 【Methods】 Two Gal4 lines (dpp-Gal4 and en-Gal4 driven by decapentaplgic and engrailed promoter, respectively, in Drosophila wing disc), afterrecombination with tub-Gal80^ts (driven by microtubulin gene tubulin promoter), were hybridized with UAS-wg (wingless) lines. The progeny larvae were cultured at a series of temperatures (18, 25, 27.5, 28, 28.5 and 30℃), and immunohistochemistry was used to detect wg transgene expression levels, which were further quantified and analyzed in the 3rd instar larvae. 【Results】 The wg transgene was not expressed at 18-30℃, but overexpressed significantly at 30℃. In the range of 25-30℃, with the increasing temperature, the wg transgene expression levels (fluorescent intensity) in en-Gal4 and dpp-Gal4 regions commonly exhibited a gradient uptrend. 【Conclusion】 The system of Gal4 and Gal80^ts combination can flexibly manipulate UAS transgene expression levels between 25-30℃. This result has an important value in the regulation of transgene expression level.    

Key words: Drosophila melanogaster , GAL4/UAS system, Gal80^ts, UAS transgene, gene expression level , fluorescence intensity