Nesvera J, Hochmannová J, Pátek M, Sroglová A, Becvárová V
Institute of Microbiology, Czech Academy of Sciences, Praha.
Appl Microbiol Biotechnol. 1994 Feb;40(6):864-6. doi: 10.1007/BF00173989.
Gram-positive facultative methylotrophic coryneform bacterium Brevibacterium methylicum was efficiently transformed with various plasmids using electroporation of intact cells. In addition to the plasmid vectors pEC71 and pZ6-1 constructed on the basis of cryptic plasmids from coryneform bacteria, broad-host-range plasmids pLS5 (derivative of plasmid pMV158 from Streptococcus agalactiae) and RSF1010 belonging to the incompatibility group IncQ from Gram-negative bacteria were found to be present as autonomous structurally unchanged DNA molecules in B. methylicum transformants. With the exception of pZ6-1, all these plasmids were stably maintained in B. methylicum cells grown under non-selective conditions. When plasmid DNAs isolated from B. methylicum were used, the highest efficiency of transformation (10(5) transformants/micrograms DNA) was achieved.
利用完整细胞电穿孔法，用各种质粒高效转化革兰氏阳性兼性甲基营养棒状杆菌甲基营养短杆菌。除了基于棒状杆菌隐蔽质粒构建的质粒载体pEC71和pZ6-1外，还发现广泛宿主范围的质粒pLS5（无乳链球菌质粒pMV158的衍生物）和属于革兰氏阴性菌不相容群IncQ的RSF1010，以自主的结构未改变的DNA分子形式存在于甲基营养短杆菌转化体中。除pZ6-1外，所有这些质粒在非选择性条件下生长的甲基营养短杆菌细胞中都能稳定维持。当使用从甲基营养短杆菌分离的质粒DNA时，实现了最高的转化效率（10⁵转化体/微克DNA）。
