Onai Kiyoshi, Morishita Megumi, Itoh Shino, Okamoto Kazuhisa, Ishiura Masahiro
Center for Gene Research, Nagoya University, Furo, Chikusa, Nagoya 464-8602, Japan.
J Bacteriol. 2004 Aug;186(15):4972-7. doi: 10.1128/JB.186.15.4972-4977.2004.
Proteins derived from the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1, which performs plant-type oxygenic photosynthesis, are suitable for biochemical, biophysical, and X-ray crystallographic studies. We developed an automated bioluminescence real-time monitoring system for the circadian clock in the thermophilic cyanobacterium T. elongatus BP-1 that uses a bacterial luciferase gene set (Xl luxAB) derived from Xenorhabdus luminescens as a bioluminescence reporter gene. A promoter region of the psbA1 gene of T. elongatus was fused to the Xl luxAB gene set and inserted into a specific targeting site in the genome of T. elongatus. The bioluminescence from the cells of the psbA1-reporting strain was measured by an automated monitoring apparatus with photomultiplier tubes. The strain exhibited the circadian rhythms of bioluminescence with a 25-h period length for at least 10 days in constant light and temperature. The rhythms were reset by light-dark cycle, and their period length was almost constant over a wide range of temperatures (30 to 60 degrees C). Theses results indicate that T. elongatus has the circadian clock that is widely temperature compensated.
来源于嗜热蓝细菌嗜热栖热放线菌BP-1(该菌进行植物型产氧光合作用)的蛋白质适用于生化、生物物理和X射线晶体学研究。我们开发了一种用于嗜热蓝细菌嗜热栖热放线菌BP-1生物钟的自动生物发光实时监测系统,该系统使用来源于发光杆菌的细菌荧光素酶基因集(Xl luxAB)作为生物发光报告基因。嗜热栖热放线菌的psbA1基因的启动子区域与Xl luxAB基因集融合,并插入到嗜热栖热放线菌基因组中的特定靶向位点。通过带有光电倍增管的自动监测装置测量psbA1报告菌株细胞的生物发光。该菌株在恒定光照和温度下,生物发光呈现出25小时周期长度的昼夜节律,至少持续10天。这些节律可通过明暗循环重置,并且在很宽的温度范围(30至60摄氏度)内其周期长度几乎恒定。这些结果表明嗜热栖热放线菌具有广泛温度补偿的生物钟。