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检测蓝藻中荧光素酶报告基因的节律性生物发光。

Detection of rhythmic bioluminescence from luciferase reporters in cyanobacteria.

作者信息

Mackey Shannon R, Ditty Jayna L, Clerico Eugenia M, Golden Susan S

机构信息

Department of Biology, Texas A&M University, College Station, USA.

出版信息

Methods Mol Biol. 2007;362:115-29. doi: 10.1007/978-1-59745-257-1_8.

Abstract

The unicellular cyanobacterium Synechococcus elongatus PCC 7942 is the model organism for studying prokaryotic circadian rhythms. Although S. elongatus does not display an easily measurable overt circadian behavior, its gene expression is under circadian control; hence, a "behavior" is created by linking a cyanobacterial promoter to either the bacterial luxAB or firefly luc luciferase genes to create reporter fusions whose activity can be easily monitored by bioluminescence. Numerous vectors have been created in our lab for introducing luciferase reporter genes into the S. elongatus chromosome. A choice of methods and equipment to detect light production from the luciferase fusions provides a means for high-throughput, automated mutant screens as well as testing rhythms from two promoter fusions within the same cell culture.

摘要

单细胞蓝藻聚球藻7942(Synechococcus elongatus PCC 7942)是研究原核生物昼夜节律的模式生物。尽管聚球藻没有表现出易于测量的明显昼夜行为,但其基因表达受昼夜节律控制;因此,通过将蓝藻启动子与细菌luxAB或萤火虫荧光素酶基因连接来创建报告融合体,从而产生一种“行为”,其活性可通过生物发光轻松监测。我们实验室已经构建了许多载体,用于将荧光素酶报告基因导入聚球藻染色体。选择检测荧光素酶融合体发光的方法和设备,为高通量、自动化突变体筛选以及在同一细胞培养物中测试两个启动子融合体的节律提供了一种手段。

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