Enríquez J A, Ramos J, Pérez-Martos A, López-Pérez M J, Montoya J
Departamento de Bioquímica y Biología Molecular y Celular, Facultad de Veterinaria, Universidad de Zaragoza, Spain.
Nucleic Acids Res. 1994 May 25;22(10):1861-5. doi: 10.1093/nar/22.10.1861.
We have developed a highly efficient DNA-synthesizing system with isolated intact rat liver mitochondria. The ATP requirements for this in organello DNA synthesis are provided by endogenous synthesis in the presence of exogenous ADP and an oxidizable substrate. In this system, mitochondrial DNA synthesis strikingly proceeds at a constant rate for about 5 h at 37 degrees C. Gel electrophoresis, hybridization and restriction enzyme analyses show that intact mitochondria synthesize nucleic acids with a size of 16.5 kb, that correspond to mitochondrial DNA, and that both DNA strands are replicated. This in organello DNA synthesis requires the supply of dNTPs and decreases at high ADP concentration in the incubation medium.
我们利用分离的完整大鼠肝线粒体开发了一种高效的DNA合成系统。该离体线粒体DNA合成所需的ATP由外源性ADP和可氧化底物存在下的内源性合成提供。在这个系统中，线粒体DNA合成在37℃下以恒定速率显著进行约5小时。凝胶电泳、杂交和限制性内切酶分析表明，完整的线粒体合成大小为16.5 kb的核酸，这与线粒体DNA相对应，并且两条DNA链都被复制。这种离体线粒体DNA合成需要供应dNTPs，并且在孵育培养基中高ADP浓度下会减少。
