Mohler W A, Blau H M
Department of Molecular Pharmacology, Stanford University Medical Center, CA 94305-5322, USA.
Proc Natl Acad Sci U S A. 1996 Oct 29;93(22):12423-7. doi: 10.1073/pnas.93.22.12423.
Complementing reporter genes provide biological indicators of coincident expression of proteins in cells. We have adapted intracistronic complementation of the Escherichia coli lacZ gene for use in mammalian cells. Enzymatic activity detectable by quantitative biochemical assay, flow cytometry, or microscopy is produced upon convergent expression of two distinct mutant lacZ peptides within single cells, or upon fusion of cells expressing such mutants. A novel fluorescent substrate for beta-galactosidase (Fluor-X-Gal) increases detection and permits simultaneous microscopic visualization of other fluorescent markers. The enzymatic complementation described here should facilitate studies of cell fusion, cell lineage, and signal transduction, by producing activity only when two proteins are expressed at the same time and place in intact cells.
互补报告基因可提供细胞中蛋白质同时表达的生物学指标。我们已将大肠杆菌lacZ基因的顺反子内互补技术应用于哺乳动物细胞。当两个不同的突变lacZ肽在单个细胞中趋同表达时，或在表达此类突变体的细胞融合时，可通过定量生化分析、流式细胞术或显微镜检测到酶活性。一种新型的β-半乳糖苷酶荧光底物（Fluor-X-Gal）提高了检测能力，并允许同时对其他荧光标记进行显微镜观察。本文所述的酶互补技术应有助于细胞融合、细胞谱系和信号转导的研究，因为只有当两种蛋白质在完整细胞的同一时间和地点表达时才会产生活性。
