Funabashi H, Imajo T, Kojima J, Kobatake E, Aizawa M
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
Luminescence. 1999 Nov-Dec;14(6):291-6. doi: 10.1002/(SICI)1522-7243(199911/12)14:6<291::AID-BIO573>3.0.CO;2-I.
The luciferase gene was introduced as a probe into a cell in order to develop a bioluminescent monitoring of intracellular ATP during fermentation. Two plasmids were constructed with two types of promoters. One was pLac-Luc, which had the luciferase gene under the lac promoter to be expressed at a high level. The other was pTet-Luc, which had the luciferase gene under the tetracycline promoter to be stably expressed. A threonine-overproducing strain of Escherichia coli (No. 29-4) was transformed with each plasmid. The recombinant E. coli strains were characterized in their growth, threonine production and luciferase expression. The bioluminescence produced intracellularly from expressed luciferase was detected during fermentation -in a non-destructive manner. The bioluminescent intensity reflected both intracellular ATP and luciferase levels, and the results indicate that stable expression of a luciferase reporter is essential for monitoring intracellular ATP.
为了在发酵过程中对细胞内ATP进行生物发光监测，将荧光素酶基因作为探针导入细胞。构建了两种带有不同类型启动子的质粒。一种是pLac-Luc，其荧光素酶基因位于lac启动子下，可高水平表达。另一种是pTet-Luc，其荧光素酶基因位于四环素启动子下，可稳定表达。用每种质粒转化一株苏氨酸高产的大肠杆菌菌株（29-4号）。对重组大肠杆菌菌株的生长、苏氨酸生产和荧光素酶表达进行了表征。在发酵过程中以非破坏性方式检测表达的荧光素酶在细胞内产生的生物发光。生物发光强度反映了细胞内ATP和荧光素酶水平，结果表明荧光素酶报告基因的稳定表达对于监测细胞内ATP至关重要。
