Lissemore J L, Jankowski J T, Thomas C B, Mascotti D P, deHaseth P L
Department of Biology, John Carroll University, University Heights, OH 44118, USA.
Biotechniques. 2000 Jan;28(1):82-4, 86, 88-9. doi: 10.2144/00281st02.
Green fluorescent protein (GFP) has become a valuable tool for the detection of gene expression in prokaryotes and eukaryotes. To evaluate its potential for quantitation of relative promoter activity in E. coli, we have compared GFP with the commonly used reporter gene lacZ, encoding beta-galactosidase. We cloned a series of previously characterized synthetic E. coli promoters into GFP and beta-galactosidase reporter vectors. Qualitative and quantitative assessments of these constructs show that (a) both reporters display similar sensitivities in cells grown on solid or liquid media and (b) GFP is especially well suited for quantitation of promoter activity in cells grown on agar. Thus, GFP provides a simple, rapid and sensitive tool for measuring relative promoter activity in intact E. coli cells.
绿色荧光蛋白(GFP)已成为检测原核生物和真核生物基因表达的一种有价值的工具。为了评估其在定量大肠杆菌中相对启动子活性方面的潜力,我们将GFP与常用的报告基因lacZ(编码β-半乳糖苷酶)进行了比较。我们将一系列先前已表征的合成大肠杆菌启动子克隆到GFP和β-半乳糖苷酶报告载体中。对这些构建体的定性和定量评估表明:(a)在固体或液体培养基上生长的细胞中,两种报告基因都表现出相似的灵敏度;(b)GFP特别适合于定量在琼脂上生长的细胞中的启动子活性。因此,GFP为测量完整大肠杆菌细胞中的相对启动子活性提供了一种简单、快速且灵敏的工具。