curing of plasmid pxo1 from bacillus anthracis using plasmid incompatibility治疗炭疽杆菌的质粒pxo1用质粒不相容.pdf

Curing of Plasmid pXO1 from Bacillus anthracis Using Plasmid Incompatibility Xiankai Liu., Dongshu Wang., Huagui Wang, Erling Feng, Li Zhu, Hengliang Wang* State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Biotechnology, Beijing, China Abstract The large plasmid pXO1 encoding the anthrax toxin is important for the virulence of Bacillus anthracis. It is essential to cure pXO1 from B. anthracis to evaluate its role in the pathogenesis of anthrax infection. Because conventional methods for curing plasmids (e.g., curing agents or growth at elevated temperatures) can induce mutations in the host chromosomal DNA, we developed a specific and reliable method to eliminate pXO1 from B. anthracis using plasmid incompatibility. Three putative replication origins of pXO1 were inserted into a temperature-sensitive plasmid to generate three incompatible plasmids. One of the three plasmids successfully eliminated the large plasmid pXO1 from B. anthracis vaccine strain A16R and wild type strain A16. These findings provided additional information about the replication/partitioning of pXO1 and demonstrated that introducing a small incompatible plasmid can generate plasmid-cured strains of B. anthracis without inducing spontaneous mutations in the host chromosome. Citation: Liu X, Wang D, Wang H, Feng E, Zhu L, et al. (2012) Curing of Plasmid pXO1 from Bacillus anthracis Using Plasmid Incompatibility. PLoS ONE 7(1): e29875. doi:10.1371/journal.pone.0029875 Editor: Adam Driks, Loyola University Medical Center, United States of America Received July 20, 2011; Accepted December 5, 2011; Published January 11, 2012 Copyright: 2012 Liu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the origina