Deciphering the physiological functions of fundamental cellular processes has until recently be hindered by the lack of molecular tools to manipulate essential genes. Recent breakthroughs in genome engineering, such as the CRISPR-Cas9 technology, have facilitated the characterization of these under-studied processes. In the frame of Eternal Cell: life without replication, we propose to develop the CRISPR-Cas9 technology for the creation of a cell that looses the ability of DNA replication during any point of the bacterial life cycle. With this approach, we intend to characterize the basic principle of DNA replication and learn more about its role for life. Using modern –omics, we will characterize the birth, life and death of these cells under normal and stress conditions and utilize our eternal cell population for applied biotechnological and medical hypotheses.