Gendel S M
Department of Genetics, Iowa State University, Ames 50011.
J Bacteriol. 1987 Oct;169(10):4426-30. doi: 10.1128/jb.169.10.4426-4430.1987.
Transposon Tn5 was used to produce random insertions in two hybrid cloning vectors for the unicellular cyanobacterium Anacystis nidulans. The transposon-containing plasmids were used to localize essential replication functions and to characterize the stability of large inserts in these vectors. The effect of the insertions on plasmid function was tested by transformation into a derivative of A. nidulans that had been cured of the endogenous plasmid used to construct the vectors. A region of approximately 4 kilobases was essential for successful plasmid transformation and replication. This region has also been shown to be involved in plasmid replication by deletion analysis. High rates of excision of Tn5 inserts within this region and restoration of normal replication function were observed when transformants were selected by using a resistance marker outside the replication region in the absence of selection for the transposon-coded kanamycin resistance. Transposon inserts outside this region were not deleted.
转座子Tn5被用于在单细胞蓝藻集胞藻6803的两种杂交克隆载体中产生随机插入。含转座子的质粒被用于定位必需的复制功能，并表征这些载体中大片段插入片段的稳定性。通过转化到已去除用于构建载体的内源质粒的集胞藻6803衍生物中，测试插入对质粒功能的影响。大约4千碱基的区域对于成功的质粒转化和复制至关重要。通过缺失分析也已表明该区域参与质粒复制。当在没有转座子编码的卡那霉素抗性选择的情况下，使用复制区域外的抗性标记选择转化子时，观察到该区域内Tn5插入片段的高切除率和正常复制功能的恢复。该区域外的转座子插入片段未被删除。
